(625f) Evaluation of Vascular Targeted Carriers Designed with Dual Ligand Strategies to Target an Inflamed Endothelium

During inflammation, activated endothelial cells (ECs) upregulate surface leukocyte adhesion molecules (LAMs) such as selectins, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1). This enables firm capture of circulating leukocytes by activated ECs and assists extravasation into the tissue to perform various functions. These same LAMs can be harnessed to localize drug delivery carriers to the diseased endothelium. Vascular-targeted carriers (VTCs) are designed as leukocyte mimics and decorated with ligands that target leukocyte adhesion molecules (LAMs) to facilitate adhesion. Due to target vessel sizes and the presence of blood flow, VTCs require different design considerations than many other targeted particle therapies; adhesion of VTCs in regions with dynamic blood flow requires multiple ligand-receptor (LR) pairs that provide particle adhesion and disease specificity. Despite the ultimate goal of leukocyte mimicry, the specificity of multiple LAM-targeted VTCs remains poorly understood, especially in physiological environments. Here, we develop a series of VTCs with well-controlled ligand properties to investigate particle binding to inflamed endothelium. Particle adhesion was evaluated in vitro to an inflamed human umbilical vein endothelial cell (HUVEC) monolayer using a parallel plate flow chamber (PPFC) and in vivo in a model of mesentery inflammation via intravital microscopy. Combining sites of sialyl Lewis A (sLe^A), a ligand for selectin, and anti-ICAM onto a single particle resulted in ~3-7-fold increase of adherent particles at the endothelium over single-ligand particles. At a constant total ligand density, a particle with a ratio of 75% sLe^A: 25% anti-ICAM resulted in more than 3-fold increase over all over other ligand ratios tested in our in vivo model. Combined with in vitro and in silico data, we find the best dual-ligand design of a particle is heavily dependent on the surface expression of the endothelial cells. Interestingly, the greatest adhesion was observed for VTCs with more ligand for the lesser-expressed EC receptor. These results establish the importance of characterizing endothelial expression associated with particular diseases and of considering LR-kinetics in intelligent VTC ligand design for future therapeutics.