Targeting of immunoliposomes to endothelial cells using a single-chain Fv fragment directed against human endoglin (CD105).

We generated immunoliposomes targeting proliferating endothelial cells by chemically coupling a single-chain Fv fragment (scFv A5) directed against human endoglin to the liposomal surface. For this purpose, we introduced an additional cysteine residue at the C-terminus of the scFv fragment. This scFv' fragment was expressed in soluble form in bacteria and allowed for a site-directed coupling to sulfhydryl-reactive lipids incorporated into the lipid bilayer. The immunoliposomes (ILA5) showed rapid and strong binding to human endoglin-expressing endothelial cells (HUVEC, HDMEC), while no binding was observed with various endoglin-negative cell lines and blood lymphocytes. In vitro, ILA5 were stable for several hours in serum- or plasma-containing medium. Incubation of endothelial cells with ILA5 at 37 degrees C led to increased binding and internalisation of the liposomes as evidenced by a perinuclear accumulation. In vitro, doxorubicin-loaded ILA5 showed an increased cytotoxicity towards endothelial cells compared to untargeted liposomes and free doxorubicin. Since the vasculature of tumours is easily accessible to drug carrier systems, the described endothelial cell-specific immunoliposomes may be useful for the development of efficacious and safe vascular targeting agents in cancer therapy.