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To assess the possibilities of retargeting adenovirus to activated endothelial cells, we conjugated bifunctional polyethylene glycol (PEG) onto the adenoviral capsid to inhibit the interaction between viral knob and coxsackie-adenovirus receptor (CAR). Subsequently, we introduced an alphav integrin-specific RGD peptide or E-selectin-specific antibody to the other functional group of the PEG molecule for the retargeting of the adenovirus to activated endothelial cells. In vitro studies showed that this approach resulted in the elimination of transgene transfer into CAR-positive cells, while at the same time specific transgene transfer to activated endothelial cells was achieved. PEGylated, retargeted adenovirus showed longer persistence in the blood circulation with area under plasma concentration-time curve (AUC) values increasing 12-fold compared to unmodified virus. Anti-E-selectin antibody-PEG-adenovirus selectively homed to inflamed skin in mice with a delayed-type hypersensitivity (DTH) inflammation, resulting in local expression of the reporter transgene luciferase. This is the first study showing the benefits of PEGylation on adenovirus behavior upon systemic administration. The approach described here can form the basis for further development of adenoviral gene therapy vectors with improved pharmacokinetics and increased efficiency and specificity of therapeutic gene transfer into endothelial cells in disease.

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In chronic inflammatory conditions, endothelial cells actively recruit immune cells from the circulation into the underlying tissue and participate in angiogenesis to support the continuous demand for oxygen and nutrients. They do so in response to activation by cytokines and growth factors such as tumour necrosis factor alpha (TNFalpha), interleukin-1 (IL-1), vascular endothelial growth factor (VEGF), and fibroblast growth factors (FGFs). Receptor triggering initiates intracellular signal transduction leading to activation of nuclear factor kappaB (NFkappaB), mitogen activated protein kinase (MAPK) activity, and nitric oxide and reactive oxygen species production, among others. As a result, adhesion molecules, cytokines and chemokines, and a variety of other genes are being expressed that mediate and control the inflammatory process. In recent years, different classes of drugs have been developed that interfere with selected enzymes involved in the intracellular signalling cascades. In endothelial cell cultures, they exert potent inhibitory effects on the expression of genes, while several studies also report on in vivo effectiveness to confine the inflammatory responses. To prevent undesired toxicity and to improve drug behaviour and efficacy, drug carrier systems have been developed that selectively deliver the therapeutics into the activated endothelial cells. The above subjects are recapitulated to give an overview on the status of development of endothelial cell directed therapeutic strategies to pharmacologically interfere with chronic inflammatory diseases.