Effects of Gamma Irradiation on the Transduction of Dividing and Nondividing Cells in Brain and Muscle of Rats by Adeno-Associated Virus Vectors

Abstract

Vectors based on adeno-associated virus (AAV) are under investigation for use in gene therapy applications. Critical aspects of AAV vector biology remain undefined, in particular the intracellular events and activities mediating transduction and determining host cell permissiveness for transduction. Using cultured primary human fibroblasts, we previously showed that AAV vectors preferentially, but not exclusively, transduce cells in the S phase of the cell cycle, and that transduction can be markedly enhanced by pretreatment of target cells with physical and chemical agents that perturb DNA metabolism. In this study, we tested whether similar improvements in AAV vector performance might be achievable in vivo. The adult rat brain and overlying scalp muscle were selected for vector inoculation because of the presence of well-defined populations of dividing, quiescent, and post-mitotic cells, and gamma irradiation was chosen as a reproducible means of inducing DNA repair in these cells. We find that gamma irradiation markedly enhances the transduction of dividing cell populations in the pia-arachnoid and choroid epithelium within the central nervous system, and of mature nondividing muscle cells in the scalp, whereas gamma irradiation did not increase the basal transduction level of post-mitotic neurons in the hippocampus. These data confirm that replicative cellular DNA synthesis is not required for transduction by AAV vectors and show that the mitotic state of target cells is not necessarily predictive of responsiveness to transduction-enhancing treatments. Most importantly, these data demonstrate that target cells can be manipulated in vivo to render them more permissive for AAV vector transduction. The transduction efficiency of adeno-associated virus (AAV) vectors varies dramatically among target cell types reflecting a critical dependence on host cell activities in the transduction process. The permissiveness of target cells for transduction by AAV vectors can be manipulated in vitro with resultant improvements in transduction efficiency of over two orders of magnitude. The data presented in this paper demonstrate that the same effect can be achieved in vivo. Combining AAV vector delivery with transduction-enhancing pretreatment of target cell populations has the potential to improve the prospects of this delivery system dramatically in gene therapy applications.