Use of a Phage Display Library to Identify Oligopeptides Binding to the Lumenal Surface of Polarized Endothelium byEx VivoPerfusion of Human Umbilical Veins

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Abstract
Human endothelial-specific targeting peptides were identified by biopanning within freshly-obtained human umbilical cords. Umbilical veins were cleaned in situ and M13 phage display libraries were passed through the cords. Tightly bound phage were recovered following isolation of endothelial cells by collagenase digestion and homogenisation, allowing production of enriched phage libraries for subsequent rounds of panning. After five rounds of biopanning, five promising sequences were selected and the binding of the corresponding phage clones was compared in perfused umbilical veins. Each of these peptides showed substantial binding, although the clone encoding the heptapeptide KPSGLTY showed the greatest, some 89-times greater than insertless phage. Binding of this phage clone was examined to cells in vitro, where it demonstrated at least five-times greater binding to isolated human umbilical vein endothelial cells than to 911, SKOV3, B16F10 and Cos7 cells. These initial peptides may prove useful targeting agents for endothelial-selective delivery, and this powerful approach should be readily applicable to biopanning in a broad range of human vessels ex vivo.