Physicochemical and biological impacts of light stress on adeno-associated virus serotype 6

Recombinant adeno-associated virus (rAAV) is a small, nonenveloped virus vector that has attracted attention as a gene therapy platform. Several studies have revealed the stability characteristics of rAAV; however, its tolerance against light stress remains unclear. Here, the physicochemical and biological impacts of light stress on rAAV6 with EGFP as a gene of interest were investigated. The results revealed that light exposure led to an approximately 20% loss in particle number of rAAV with s value of full particles, whereas a more drastic decrease was observed in the genomic titer due to the degradation of encapsidated DNA, leading to an approximately 90% reduction in biological activity as evident from a flow cytometry-based assay. Structure analysis of the viral proteins VP1, VP2, and VP3 and the encapsidated DNA revealed that light exposure causes typical photodegradation of protein and DNA, but with AAV-specific phenomena. This included oxidation proceeding at histidine residues of VP3 on the inner surface of the capsid and the formation of cyclobutane pyrimidine dimer in DNA within the capsid. The findings highlight the necessity of the proper handling and storage to maintain the stability and efficacy of AAV-based gene therapies.