Targeting polymer coated adenovirus to tumour-associated vasculature

Thesis

Tumour-associated vasculature provides an accessible target for systemic gene therapy using targeted adenoviruses. The aim of this thesis is to develop strategies for targeting adenovirus infection to tumour-associated endothelium.

Adenovirus expressing luciferase (Adluc) was coated with an amino-reactive polymer based on poly [N-(2-hydroxypropyl) methacrylamide] [pHPMA] to ablate normal infection pathways¬. This was a pre-requisite to redirecting virus tropism to infect endothelial cells via specific receptors.

Direct attachment to the pHPMA-adenovirus (pcAdluc) of ligands including vascular endothelial growth factor (VEGF165) and a monoclonal antibody (RAFL) recognising VEGF receptor 2 (VEGFR-2) retargeted infectivity to VEGFR-2-positive endothelial cells and not to receptor-negative cells. Specificity of transduction in vitro was shown by competition with excess antibody. In vivo however, the VEGF165-retargeted virus failed to transduce tumour-associated endothelia following systemic administration. Similarly, direct linkage of a monoclonal antibody against E-Selectin (MHES) demonstrated E-Selectin-specific transduction of tumour necrosis factor-α (TNF-α)-activated endothelial cells, although overall levels of infection were not increased compared to unmodified Adluc.

A two-component targeting system using protein A or protein G as ‘bridging’ agents was developed to ensure the required orientation of targeting antibodies. Using this system MHES mediated greater transduction of TNF-α-activated endothelial cells than Adluc. Conjugation using protein A also gave non-specific effects which were not seen with protein G. Whereas the unmodified Adluc virus failed to transduce TNF-α-activated endothelium in an umbilical vein model ex vivo, the MHES-protein G-pHPMA-adenovirus (MHES-StrepGpcAdluc) mediated good transduction. Similarly, StrepGpcAdluc retargeted with a chimeric P-Selectin Glycoprotein Ligand-1 (PSGL-1)-Fc fusion protein, showed good circulation kinetics and significant uptake into HepG2 xenografts following intravenous administration. Histological studies suggested selective targeting to tumour-associated endothelial cells.

Overall these findings support the assertion that tumour-associated vasculature is an accessible target for systemic gene delivery, and the use of protein G as bridging agent facilitates rapid screening of Fc-bearing ligands for retargeting pcAd infection to tumour-associated endothelium.

Authors: Bachtarzi, H

• Publication date: 2010
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: pHPMA; VEGFR-2; adenovirus; E-Selectin; cancer; vascular targeting; gene delivery
• Subjects: Tumours; Oncology; Vascular research; Biology (medical sciences); Viruses; Medical Sciences; Gene medicine; Nano-biotechnology; Life Sciences; Pharmacology