Improved Animal Models for Testing Gene Therapy for Atherosclerosis

Abstract

Gene therapy delivered to the blood vessel wall could augment current therapies for atherosclerosis, including systemic drug therapy and stenting. However, identification of clinically useful vectors and effective therapeutic transgenes remains at the preclinical stage. Identification of effective vectors and transgenes would be accelerated by availability of animal models that allow practical and expeditious testing of vessel-wall-directed gene therapy. Such models would include humanlike lesions that develop rapidly in vessels that are amenable to efficient gene delivery. Moreover, because human atherosclerosis develops in normal vessels, gene therapy that prevents atherosclerosis is most logically tested in relatively normal arteries. Similarly, gene therapy that causes atherosclerosis regression requires gene delivery to an existing lesion. Here we report development of three new rabbit models for testing vessel-wall-directed gene therapy that either prevents or reverses atherosclerosis. Carotid artery intimal lesions in these new models develop within 2-7 months after initiation of a high-fat diet and are 20-80 times larger than lesions in a model we described previously. Individual models allow generation of lesions that are relatively rich in either macrophages or smooth muscle cells, permitting testing of gene therapy strategies targeted at either cell type. Two of the models include gene delivery to essentially normal arteries and will be useful for identifying strategies that prevent lesion development. The third model generates lesions rapidly in vector-naïve animals and can be used for testing gene therapy that promotes lesion regression. These models are optimized for testing helper-dependent adenovirus (HDAd)-mediated gene therapy; however, they could be easily adapted for testing of other vectors or of different types of molecular therapies, delivered directly to the blood vessel wall. Our data also supports the promise of HDAd to deliver long-term therapy from vascular endothelium without accelerating atherosclerotic disease.

Local Vascular Gene Therapy With Apolipoprotein A-I to Promote Regression of Atherosclerosis

“…As expected, 34 intimal lesions were rich in lipid and macrophages (Figure 3B, 3C, 3F, and 3G). The lesions also contained smooth muscle cells, typically in a lesion cap (Figure 3D and 3H) and rare T cells (Figure V in the online-only Data Supplement).…”

“…We measured plasma cholesterol after 1 month of this diet and then every two weeks. As expected, 34 rabbits had a heterogeneous response to 1 month of fat-feeding, with plasma cholesterols ranging from 200 mg/dL to over 2,000 mg/dL. Subsequent adjustment of dietary cholesterol, according to a sliding scale, narrowed the range of plasma cholesterols substantially, and maintained the majority of rabbits in the targeted 200–800 mg/dL range during the period of lesion progression (4 weeks–28 weeks).…”

“…These historical control arteries were from rabbits that also underwent bilateral carotid infusions of DMEM after 4 weeks of high-fat diet but were harvested at the 28-week time point instead of receiving infusions of HDAd. 34 The smaller intimal areas in the present study suggest that our protocol of diet-induced lowering of plasma cholesterol promoted regression of atherosclerosis in both groups. Consistent with findings of the study in which we developed this model, 34 lesion size was highly variable with intimal areas ranging from 355 μm 2 to 550,000 μm 2 in individual arteries.…”

“…34 The smaller intimal areas in the present study suggest that our protocol of diet-induced lowering of plasma cholesterol promoted regression of atherosclerosis in both groups. Consistent with findings of the study in which we developed this model, 34 lesion size was highly variable with intimal areas ranging from 355 μm 2 to 550,000 μm 2 in individual arteries. This variability, along with the high correlation of intimal areas between left and right arteries in the same rabbit (r=0.7; P <0.0002; Figure IV in the online-only Data Supplement), supported our prospective decision to use a paired sample study design for the final data analysis.…”

“…To test whether vascular apo A-I gene therapy promotes atherosclerosis regression, we first developed a rabbit model of focal severe carotid atherosclerosis. 34 This model yields large intimal lesions that are macrophage and lipid-rich, but also contain smooth muscle cells, often in a fibrous cap. These lesions resemble mature human plaques and are—as noted by others 46 —a more clinically relevant target than the early fatty streaks present in many mouse models and in our rabbit model of atherosclerosis progression.…”