ICAM-directed vascular immunotargeting of antithrombotic agents to the endothelial luminal surface.

Drug targeting to a highly expressed, noninternalizable determinant up-regulated on the perturbed endothelium may help to manage inflammation and thrombosis. We tested whether inter-cellular adhesion molecule-1 (ICAM-1) targeting is suitable to deliver antithrombotic drugs to the pulmonary vascular lumen. ICAM-1 antibodies bind to the surface of endothelial cells in culture, in perfused lungs, and in vivo. Proinflammatory cytokines enhance anti-ICAM binding to the endothelium without inducing internalization. (125)I-labeled anti-ICAM and a reporter enzyme (beta-Gal) conjugated to anti-ICAM bind to endothelium and accumulate in the lungs after intravenous administration in rats and mice. Anti-ICAM is seen to localize predominantly on the luminal surface of the pulmonary endothelium by electron microscopy. We studied the pharmacological effect of ICAM-directed targeting of tissue-type plasminogen activator (tPA). Anti-ICAM/tPA, but not control IgG/tPA, conjugate accumulates in the rat lungs, where it exerts plasminogen activator activity and dissolves fibrin microemboli. Therefore, ICAM may serve as a target for drug delivery to endothelium, for example, for pulmonary thromboprophylaxis. Enhanced drug delivery to sites of inflammation and the potential anti-inflammatory effect of blocking ICAM-1 may enhance the benefit of this targeting strategy.

Platelet-endothelial cell adhesion molecule-1-directed immunotargeting to cardiopulmonary vasculature.

Therapeutic molecules conjugated with antibodies to the platelet-endothelial cell adhesion molecule-1 (PECAM-1) accumulate in the pulmonary endothelium after i.v. injection in mice. In this study, we characterized PECAM-directed targeting to the lung and heart after local versus systemic intravascular administration in a large animal model, pigs. Radiolabel tracing showed that 1 h post-i.v. injection, 35% of anti-PECAM versus 2.5% of control IgG had accumulated in the lungs. Infusion of anti-PECAM via a catheter placed in the right pulmonary artery (RPA) resulted in a 3-fold elevation of the uptake in the right lower lobe and 2-fold reduction of uptake in the left lobes in the lung. Cardiac uptake of anti-PECAM was negligible after i.v. and RPA infusion. In contrast, delivery with a catheter placed in the right coronary artery (RCA) resulted in a 4-fold elevation of cardiac uptake of anti-PECAM, but not IgG, compared with i.v. injection. To estimate the targeting of an active reporter enzyme, streptavidin-conjugated beta-galactosidase (beta-Gal) was coupled to anti-PECAM or IgG (anti-PECAM/beta-Gal and IgG/beta-Gal) and injected into the RCA. Beta-Gal activity was markedly elevated in the heart and lungs (5- and 25-fold increased, respectively) after injection of anti-PECAM/beta-Gal, but not IgG/beta-Gal. Image analysis confirmed endothelial targeting of anti-PECAM/beta-Gal in the heart and lung. In summary, anti-PECAM antibody conjugates deliver agents to the pulmonary endothelium regardless of injection route, whereas local arterial infusion permits targeting to the cardiac vasculature. This paradigm may be useful for drug targeting to endothelium in lungs, heart, and possibly other organs.

Platelets inhibit the lysis of pulmonary microemboli.

Using tracings of (125)I-labeled fibrin(ogen) in rodents, we examined the hypothesis that platelets impede the lysis of pulmonary emboli. (125)I-Microemboli (ME, 3-10 micron diameter) lodged homogeneously throughout the lungs after intravenous injection in both rats and mice (60% of injected dose), caused no lethality, and underwent spontaneous dissolution (50 and 100% within 1 and 5 h, respectively). Although lung homogenates displayed the most intense fibrinolytic activity of all the major organs, dissolution of ME was much slower in isolated perfused lungs (IPL) than was observed in vivo. Addition of rat plasma to the perfusate facilitated ME dissolution in IPL to a greater extent than did addition of tissue-type plasminogen activator alone, suggesting that permeation of the clot by plasminogen is the rate-limited step in lysis. Platelet-containing ME injected in rats lysed much more slowly than did ME formed from fibrin alone. (125)I-Thrombi, formed in the pulmonary vasculature of mice in response to intravascular activation of platelets by injection of collagen and epinephrine, were essentially resistant to spontaneous dissolution. Moreover, injection of the antiplatelet glycoprotein IIb/IIIa antibody 7E3 F(ab')(2) facilitated spontaneous dissolution of pulmonary ME and augmented fibrinolysis by a marginally effective dose of Retavase (10 microg/kg) in rats. These studies show that platelets suppress pulmonary fibrinolysis. The mechanism(s) by which platelets stabilize ME and utility of platelet inhibitors to facilitate their dissolution deserves further study.