Incorporation of tumor vasculature targeting motifs into moloney murine leukemia virus Env escort proteins enhances retrovirus binding and transduction of human endothelial cells

Adhesion receptors expressed on the surfaces of tumor-activated endothelial cells provide an advantageous locus for targeting gene therapy vectors to angiogenic tissues and/or tumor vasculature. In this study, we engineered a series of Bsn-Gly-Arg (NGR)-containing congeners of the presumptive cell b inding motif contained within the ninth type III repeat of fibronectin and displayed these tumor vasculature targeting motifs (TVTMs) within the conte xt of Moloney murine leukemia envelope "escort" proteins. Comparative studi es of envelope incorporation into viral particles and evaluation of the cel l binding properties of the targeted vectors revealed critical structural f eatures, thus identifying a subset of optimal TVTMs. Utilizing a modified E LISA. to evaluate viral binding to target cells, we observed a significant down-regulation of TVTM-virion binding to human endothelial cells following sustained (48-h) exposure to VEGF. Normalized for equivalent titers (10(6) CFU/ml), as assayed on NIH 3T3 cells, vectors displaying TVTM escort prote ins significantly enhanced the transduction efficiency from 12.2 to 37.4% i n human KSY-1 endothelial cell cultures (P < 0.001) and from 0.4 to 4.1% in human umbilical vein endothelial cell (HUVEC) cultures (P < 0.001). In sum mary, these studies utilized an engineering approach to identify a subset o f TVTMs that are stably incorporated as envelope ''escort" proteins into re troviral vectors and that, by functioning to improve the binding efficiency and transduction of both HUVEC and KSY1 endothelial cells, may have therap eutic potential for targeting gene delivery to the tumor-associated vascula ture,