Platelet-Endothelial Cell Adhesion Molecule-1-Directed Immunotargeting to Cardiopulmonary Vasculature

Scherpereel, Arnaud; Rome, Jonathan J.; Wiewrodt, Rainer; Watkins, Simon C.; Harshaw, David W.; Alder, S A; Christofidou‐Solomidou, Melpo; Haut, Elliott R.; Murciano, Juan-Carlos; Nakada, Marian T.; Albelda, Steven Μ.; Muzykantov, Vladimir R.

doi:10.1124/jpet.300.3.777

Cited by 60 publications

(80 citation statements)

References 36 publications

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“…Understanding the mechanisms that regulate uptake of anti-ICAM-1 and anti-PECAM-1 conjugates will probably help to extend the utility of these agents as the basis for endotheliumspecific drug-targeting vehicles (Li et al, 2000;Muzykantov et al, 1999;Scherpereel et al, 2002;Scherpereel et al, 2001;Wiewrodt et al, 2002). For instance, inhibitors of conjugate uptake might help to increase their stability by reducing the extent of delivery to lysosomes and other degradative compartments.…”

Section: Discussionmentioning

confidence: 99%

A novel endocytic pathway induced by clustering endothelial ICAM-1 or PECAM-1

Muro

Wiewrodt

Thomas

et al. 2003

Journal of Cell Science

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273

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Antibody conjugates directed against intercellular adhesion molecule(ICAM-1) or platelet-endothelial cell adhesion molecule (PECAM-1) have formed the basis for drug delivery vehicles that are specifically recognized and internalized by endothelial cells. There is increasing evidence that ICAM-1 and PECAM-1 may also play a role in cell scavenger functions and pathogen entry. To define the mechanisms that regulate ICAM-1 and PECAM-1 internalization, we examined the uptake of anti-PECAM-1 and anti-ICAM-1 conjugates by endothelial cells. We found that the conjugates must be multimeric, because monomeric anti-ICAM-1 and anti-PECAM-1 are not internalized. Newly internalized anti-ICAM-1 and anti-PECAM-1 conjugates did not colocalize with either clathrin or caveolin, and immunoconjugate internalization was not reduced by inhibitors of clathrin-mediated or caveolar endocytosis, suggesting that this is a novel endocytic pathway. Amiloride and protein kinase C (PKC) inhibitors, agents known to inhibit macropinocytosis,reduced the internalization of clustered ICAM-1 and PECAM-1. However,expression of dominant-negative dynamin-2 constructs inhibited uptake of clustered ICAM-1. Binding of anti-ICAM-1 conjugates stimulated the formation of actin stress fibers by human umbilical vein endothelial cells (HUVEC). Latrunculin, radicicol and Y27632 also inhibited internalization of clustered ICAM-1, suggesting that actin rearrangements requiring Src kinase and Rho kinase (ROCK) were required for internalization. Interestingly, these kinases are part of the signal transduction pathways that are activated when circulating leukocytes engage endothelial cell adhesion molecules, suggesting the possibility that CAM-mediated endocytosis is regulated using comparable signaling pathways.

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Section: Discussionmentioning

confidence: 99%

A novel endocytic pathway induced by clustering endothelial ICAM-1 or PECAM-1

Muro

Wiewrodt

Thomas

et al. 2003

Journal of Cell Science

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273

525

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“…However, prophylactic fibrinolysis by PECAM-targeted PA may help to prevent extension/recurrence of pulmonary thromboembolism 32,46 and attenuate acute lung graft rejection because of thrombotic occlusion of the pulmonary vasculature. 47 Because of the pan-endothelial nature of PECAM expression and the opportunity to enrich local delivery using catheters, 17 a wider utility to protect other vascular beds against imminent thrombosis can be envisioned. The cardiac accumulation of anti-PECAM scFv-uPA noted in this work ( Figure 4A) is of interest in this context.…”

Section: Discussionmentioning

confidence: 99%

“…14,15 These molecules are especially attractive targets for delivering antithrombotic agents, because (1) they are expressed by endothelial cells and are actively involved in sites of inflammation and thrombosis 16 ; (2) after intravenous injection, anti-PECAM and anti-ICAM conjugates accumulate in the pulmonary vasculature, a common site of thromboembolism; (3) these conjugates can also deliver drugs to endothelium in other organs, such as, in cardiac or cerebral vasculature, through infusion via their respective afferent arteries 17 ; and (4) endothelium does not internalize anti-PECAM and anti-ICAM, allowing antithrombotic activity to be maintained on the luminal surface. 14,18 We have also found, however, that multivalent conjugates, formed by chemical crosslinking of antibodies and drugs, cluster endothelial PECAM or ICAM, leading to endocytosis and lysosomal degradation, 16,19 which terminates the effectiveness of delivered enzymes despite high activity and successful targeting in vivo.…”

Section: Introductionmentioning

confidence: 99%

Endothelial targeting of a recombinant construct fusing a PECAM-1 single-chain variable antibody fragment (scFv) with prourokinase facilitates prophylactic thrombolysis in the pulmonary vasculature

Ding

Gottstein

Grunow

et al. 2005

Blood

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Means to prevent thrombus extension and local recurrence remain suboptimal, in part because of the limited effectiveness of existing thrombolytics. In theory, plasminogen activators could be used for this purpose if they could be anchored to the vascular lumen by targeting stably expressed, noninternalized determinants such as platelet-endothelial-cell adhesion molecule 1 (PECAM-1). We designed a recombinant molecule fusing lowmolecular-weight single-chain prourokinase plasminogen activator (lmw-scuPA) with a single-chain variable fragment (scFv) of a PECAM-1 antibody to generate the prodrug scFv/lmw-scuPA. Cleavage by plasmin generated fibrinolytically active 2-chain lmw-uPA. This fusion protein (1) bound specifically to PECAM-1-expressing cells; (2) was rapidly cleared from blood after intravenous injection; (3) accumulated in the lungs of wild-type C57BL6/J, but not PECAM-1 null mice; and (4) lysed pulmonary emboli formed subsequently more effectively than lmw-scuPA, thereby providing support for the concept of thromboprophylaxis using recombinant scFv-fibrinolytic fusion proteins that target endothelium. ( IntroductionPlasminogen activators (PAs; eg, uPA, urokinase plasminogen activator) help to restore perfusion after thrombotic vascular occlusion, the leading cause of human morbidity and mortality. [1][2][3] However, the clinical utility of PAs is limited by (1) inadequate delivery because of rapid elimination and inactivation en route and ineffective penetration into formed clots; (2) side effects, including extravasation leading to collateral damage in the central nervous system and other tissues; (3) lysis of "physiologic" (hemostatic) clots leading to hemorrhage; and, (4) reperfusion injury following a delay in restoring perfusion, where morbidity correlates with the duration of ischemia. [4][5][6] Clinical settings characterized by a high propensity for thrombosis have been identified, and means to diagnose early clot formation have been developed. 1,2 Although the indications for prophylaxis are known, PAs are not used prophylactically because of their unfavorable pharmacokinetics and side effects. Gene therapy approaches, effective in cell-culture and animal experiments, 7,8 are not practical when the need to enhance fibrinolysis is acute and of short duration. 9 Conceivably, prophylactic delivery of a PA derivative that rapidly restricts and sustains its activity in the vascular lumen can help to lyse nascent clots expeditiously, inhibit propagation of mural thrombi, and reduce the duration of ischemia.For example, PAs can be used for thromboprophylaxis by coupling to carrier red blood cells (RBCs), prolonging circulation and limiting extravasation. 10 This approach may have utility in settings in which RBC transfusion is part of current management. Drug targeting to suitable endothelial-cell-surface determinants 11-13 may provide an alternative approach and, in theory, localize PA activity in the affected intravascular compartment.For example, drugs coupled with antibodies to plateletendothelial ...

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“…CytoTox 96 NonRadioactive Cytotoxicity Assay was purchased from Promega (Madison, WI). Anti-PECAM mAb 62, a monoclonal mouse IgG 2␣ antibody directed against human platelet-endothelial cell adhesion molecule-1, has been characterized in our previous studies (Muzykantov et al, 1999;Scherpereel et al, 2002). All cell culture medium components were from Life Technologies (Gaithersburg, MD) unless otherwise noted.…”

Section: Methodsmentioning

confidence: 99%

“…A particularly good EC molecule for antibody-targeting is platelet-endothelial cell adhesion molecule (PECAM)-1 (CD31), a molecule that is constitutively and stably expressed on the endothelial lumen at the level of approximately a million copies per cell and is involved in transcytosis of activated leukocytes via endothelium in inflammation (Scherpereel et al, 2002). Conjugation of diverse therapeutic cargoes and carriers with anti-PECAM antibodies provides robust intracellular drug delivery into endothelium (Li et al, 2000;Muro et al, 2003b).…”

mentioning

confidence: 99%

Platelet-Endothelial Cell Adhesion Molecule-1-Directed Endothelial Targeting of Superoxide Dismutase Alleviates Oxidative Stress Caused by Either Extracellular or Intracellular Superoxide

Shuvaev¹,

Tliba²,

Nakada³

et al. 2007

J Pharmacol Exp Ther

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ABSTRACT. However, another reactive oxygen species, superoxide anion, is also involved in many forms of vascular oxidative stress, including ischemia/reperfusion, hypertension, and inflammation. To protect endothelium against superoxide attack, we designed and tested antibody-directed targeting of superoxide dismutase (SOD) to the endothelial surface determinant, platelet-endothelial cell adhesion molecule (PECAM)-1. We synthesized anti-PECAM/SOD conjugates that retained 70% of enzymatic activity (superoxide anion dismutation) and specifically bound to endothelial cells, but not PECAM-negative cells. The effect of anti-PECAM/SOD delivery to cells was tested in two distinct models of oxidative stress induced by either extracellular or intracellular generation of superoxide anion. In the first model, anti-PECAM/SOD, but not unconjugated SOD, protected endothelial cells against injury caused by superoxide produced in the medium by hypoxanthine-xanthine oxidase. At the optimal dose, anti-PECAM/SOD provided up to 40 to 50% protection against cell death in this model. In the second model, anti-PECAM/SOD at the optimal dose provided complete protection against necrosis caused by paraquat-induced intracellular superoxide generation. Endothelial targeting of SOD represents a new molecular antioxidant approach that could be used for the management of vascular oxidative stress.

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Platelet-Endothelial Cell Adhesion Molecule-1-Directed Immunotargeting to Cardiopulmonary Vasculature

Cited by 60 publications

References 36 publications

A novel endocytic pathway induced by clustering endothelial ICAM-1 or PECAM-1

A novel endocytic pathway induced by clustering endothelial ICAM-1 or PECAM-1

Endothelial targeting of a recombinant construct fusing a PECAM-1 single-chain variable antibody fragment (scFv) with prourokinase facilitates prophylactic thrombolysis in the pulmonary vasculature

Platelet-Endothelial Cell Adhesion Molecule-1-Directed Endothelial Targeting of Superoxide Dismutase Alleviates Oxidative Stress Caused by Either Extracellular or Intracellular Superoxide

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