There is much we do not yet know of drug-eluting stents and local vascular drug delivery. Questions remain as to when and why these devices function or potentially generate morbidity risk. There is not a clear understanding of how such devices function in acute thrombosis, chronic metabolic derangements like diabetes mellitus or vascular beds other than the coronary arteries. The literature suggests that efficacy of drug-eluting-stents is impacted by lesion complexity and degree of atherosclerosis (14
). Similarly, emerging data infer that drug-eluting balloons can provide significant benefit to peripheral arterial disease when introduced at the time of direct intervention on existing complex lesions(37
). The very efficacy of paclitaxel and sirolimus following local delivery is usually attributed to their lipophilicity(10
) and sustained retention in the vessel wall compared to more hydrophilic compounds like heparin(21
). It is hypothesized that deposition of lipophilic drugs will increase with arterial wall lipid content(14
) and that drug effect should track with lesion composition and morphology. Yet, the bulk of preclinical studies to date have utilized intact arteries and normal animals and many of the postulates regarding tissue deposition have not been formally tested. The current study correlated drug distribution with local arterial composition in human autopsy samples and controlled animal models of arterial disease and injury and defied this hypothesis.
The distribution of three clinically relevant hydrophobic drugs in human autopsy samples revealed changes in drug distribution with lesion state, but in a manner that cannot be explained solely by drug lipophilicity or directly with arterial wall lipid content. Remarkably, although all three drugs are hydrophobic, their compartmental deposition in the chronic atheromatous domains of the human aorta scaled inversely with compartmental cholesterol content (; p<0.0001, R=0.81). Fresh calf carotid arteries had lower levels of cholesterol than the media of the human aorta samples, and correspondingly higher drug partition coefficients (supplemental figure S3
More intricate effects were observed in controlled rabbit models that examined the compounded effects of diet and denudation on drug distribution following sustained drug incubation. The equilibrium deposition of paclitaxel and sirolimus-like drugs are differentially affected by lesion complexity. Whereas everolimus distribution in arteries that were injured at low catheter inflation volumes (0.5cc) was insensitive to differences in diet, paclitaxel distribution was significantly altered in animals that received a cholesterol rich diet (), particularly in the subinitmal region. High levels of paclitaxel in the subintimal space of mildly injured arteries correlate with a diet-induced upregulation of tubulin in that area (supplemental Fig S4
). Conversely, the apparent insensitivity of FKBP-12 distribution in mildly injured arteries to differences in diet (supplemental Fig S4
) correlated with insignificant alterations in the distribution of sirolimus (). Catheter injury ar the higher inflation volume (1.0cc) allowed us to examine the correlation of paclitaxel distribution with lesion morphology and composition in the setting of greater vascular injury and ultimate tissue response. Acute disruption following local tissue damage removes natural transport barriers that hinder the accumulation of interstitial lipid, and induces an inflammatory stimulus that allows for marked increase in local accumulation of macrophages and dendritic cells(38
). Levels of tubulin rise in injured arteries where hypercholesterolemia increases macrophage infiltration(42
) and as suspected paclitaxel deposition increases in these local areas as well. Yet, there is also a reverse effect if interstitial lipid pools are dominant in place of macrophage infiltration. Lipid pools displace tubulin expressing cells in the intima and media, thereby removing a binding domain for paclitaxel ( and ), reducing its arterial deposition in a manner that scales inversely with lipid content (). Notably, although tubulin expression was upregulated in the group of acutely injured arteries, diet abolished this effect (), speaking to the reported differences in tubulin distribution.
Thus, it is only when binding to drug-specific tissue sites are added to transport considerations(44
) that one can account for the differential deposition and distribution of drugs of near identical molecular weight, similar lipophilicity and solubility across similar arterial tissue. The differences in the dependence of drug deposition on tissue state may well represent the different balance each drug achieves between increased absorption of drug within macrophages and decreased binding in settings of lipid infiltration and cell displacement(42
). Paclitaxel, by virtue of its effects on tubulin, effectively fixes macrophages in place(10
) eliciting a mechanism for a cascade of injury, altered tissue state and affected local drug retention and perhaps effect. In contrast, sirolimus analogs were virtually unaffected by vascular manipulations (), consistent with uniform, though low, expression of FKBP-12 in a range of arteries and transient upregulation of FKBP-12 that peaks early after and returns to baseline levels late after arterial injury(41
). Intriguingly, macrophage infiltration does not chronically upregulate FKBP-12, suggesting a mechanism for differential effects of lesion complexity on the distribution and efficacy of paclitaxel and sirolimus analogs(14
). While drug binding to specific intracellular targets is important, our finding of paclitaxel colocalization with elastin (), suggests that elastin displays a high binding capacity for paclitaxel, speaking to the importance of the extracellular matrix as a determinant of the distribution and retention of small hydrophobic drugs. In vitro imaging studies with tissue mimics also illustrated colocalization of fluorescent paclitaxel with elastin, and implicated the latter as a prime drug-binding substrate that impedes paclitaxel diffusion, rather than through steric hindrance(48
The idea that drug deposition after balloon inflation and stent implantation within diseased, atheromatous and sclerotic vessels tracks so precisely with specific tissue elements is an important consideration of drug-eluting technologies and may well require that we consider diseased rather than naïve tissues in preclinical evaluations. We must acknowledge that excised and autopsy specimens might undergo structural changes that we could not see after histological characterization, and that there are ultrastructural differences and different pathophysiologic consequences of disease in abdominal aorta and coronary arteries and between human and leporine tissues. Our use of abdominal aorta from human autopsy samples and rabbits subject to controlled diet and injury, rather than coronary arteries, ensured greater tissue preservation and allowed for comparison of like tissues in best preserved state. The immersion of tissues required for observing the differences we cite are not identical with drug elution from endovascular balloons, stents or perivascular wraps that specifically target a single aspect of the artery; immersion of tissue segments in binding medium allows for drug absorption not only from the intima and adventitia but also by lateral diffusion along the tunica layers. Nevertheless, the equilibrium effects that we report are essentially independent of such transport issues and are primarily a reflection of the tissue’s equilibrium binding capacity for the drug.
The idea that the artery as a target tissue determines and regulates uptake of locally delivered drug is biologically appealing and consistent with concern raised as to the validity of evaluation of devices and drug-elution in preclinical animal models that employ normal blood vessels(14
). Though animal models cannot predict human efficacy they can be used to test mechanism of action(49
). When uninjured animal vessels are examined the extrapolation of mechanism to the clinical condition may be limited. The change in drug uptake and retention with tissue architecture and disease can begin to explain seemingly disparate findings from different clinical trials(15
). It is only when drug binding to specific tissue sites is added to transport considerations(44
) that one can account for the differential deposition and distribution of drugs of near identical molecular weight, similar lipophilicity and solubility across similar arterial tissue. Binding in turn requires an understanding of the kinetics of tissue response to injury. Indeed, the specific targets of the leading drugs eluted from stents, paclitaxel and sirolimus analogs, may express more abundantly in recruited inflammatory cells than in the native artery itself. Thus, the reaction of an artery first to the initial injury, then to the vascular repair and finally to the very effect of eluted drug will in turn influence drug absorption and distribution. It is in this way that different drugs can be absorbed by the same artery differently even at identical degrees of injury, cell infiltration and lipid insudation.