MicroRNA-based recombinant AAV vector assembly improves efficiency of suicide gene transfer in a murine model of lymphoma.

Recent success in clinical trials with recombinant Adeno-associated virus (AAV)-based gene therapy has redirected efforts in optimizing AAV assembly and production, to improve its potency. We reasoned that inclusion of a small RNA during vector assembly, which specifically alters the phosphorylation status of the packaging cells may be beneficial. We thus employed microRNAs (miR-431, miR-636) identified by their ability to bind AAV genome and also dysregulate Mitogen-activated protein kinase (MAPK) signaling during vector production, by a global transcriptome study in producer cells. A modified vector assembly protocol incorporating a plasmid encoding these microRNAs was developed. AAV2 vectors packaged in the presence of microRNA demonstrated an improved gene transfer potency by 3.7-fold, in vitro. Furthermore, AAV6 serotype vectors encoding an inducible caspase 9 suicide gene, packaged in the presence of miR-636, showed a significant tumor regression (~2.2-fold, P < .01) in a syngeneic murine model of T-cell lymphoma. Taken together, we have demonstrated a simple but effective microRNA-based approach to improve the assembly and potency of suicide gene therapy with AAV vectors.