Stability of Recombinant Mosaic Adeno-Associated Virus Vector rAAV/DJ/CAG at Different Temperature Conditions

Abstract

The nanometer size and biological characteristics of recombinant adeno-associated virus vectors (rAAV) make them particularly useful as gene therapy vectors and they have been successfully used in this role. Our latest research revealed that the rAAV/DJ/CAG mosaic vector offers highly efficient targeted gene delivery to melanoma cells metastasized to the lungs and that the transduction is temperature dependent. In order to further explore the ability of the rAAV/DJ/CAG vector to deliver highly selective transduction, this study was designed to identify the transduction stability of rAAV/DJ/CAG under various conditions. The temperatures used in this study ranged from −196 ° (liquid nitrogen) to 90 °, and the effect of temperature fluctuations (freeze-thaw, cooling-heating cycles) was also studied. This research also investigated the effects of UV radiation (ultraviolet) on the rAAV/DJ/CAG activity. Changes in the transduction efficiency were assessed via fluorescence microscopy imaging and the qPCR method. Under the test conditions, the transduction efficiency was reduced by approx. 35%, on average. High temperatures (70 °/90 °) and UV light proved to have the most detrimental impact. Changes in the stability of the rAAV/DJ/CAG structure are manifested by variations in the number of genome copies (gc) and GFP+ cells. Temperature fluctuations resulted in differences in the number of gc while maintaining a similar number of GFP+ cells, which may indicate specific changes in the rAAV/DJ/CAG structure, triggering disorders or degradation in the vector entry. This study provides interesting insights into rAAV/DJ/CAG, and the implications of these findings provide a basis for developing new protocols in cancer gene therapy.