In the high-stakes game of gene therapy, adeno-associated virus (AAV) vectors have been the go-to workhorses. These vectors have the ability to deliver a therapeutic gene to a patient’s cells, offering hope for the treatment of genetic disorders, such as Duchenne muscular dystrophy, hemophilia A, and certain retinal degeneration disorders. But, akin to a mail carrier trying to stuff an oversized package into a small mailbox, AAV vectors have been stymied by their limited packaging capacity for sizable therapeutic transgenes.
Enter the exciting, albeit technically challenging, realm of dual-vector AAV systems. The notion is akin to the strategy parcel delivery services use when a package is too large for one box, they split the contents into two boxes. Scientists are now working to divide larger transgenes between two AAV vectors, aiming to overcome the limitations of single-vector systems and potentially opening the door to the treatment of disorders caused by big gene mutations.
The need for such an approach cannot be overstated. While retroviruses have the room to accommodate larger transgenes, they are like a double-edged sword. Their use can be fraught with greater risks due to potential insertional mutagenesis or oncogene activation – an equivalent of delivering a package that unintentionally disrupts the household and may even start a fire. Lentiviral vectors, on the other hand, offer some promise but are like a courier that can’t find the address, falling short in terms of safety and efficacy compared to AAV, particularly when it comes to transducing non-dividing cells of the central nervous system.
The development of dual-vector AAV systems is not a straightforward task. It’s not as simple as splitting a transgene in half and placing the halves into separate vectors. The process involves the creation of two AAV vectors, each carrying a piece of the oversized transgene. Upon delivery to the host cell, the transgene parts must find each other, recombine, and function as intended. This could occur due to single-strand annealing of plus and minus strands at the region of homology or by homologous recombination (HR) following second-strand synthesis of the truncated transgenes.
However, there are hurdles to this approach. Fragmented packaging of oversized transgenes can lead to different outcomes, including non-homologous end joining (NHEJ) of transgenes following second-strand synthesis, which may also occur in combination with ITR concatemerization.
As the industry continues to push the boundaries of what’s possible in gene therapy, the development of dual-vector AAV systems represents an important frontier. It’s a scientific gamble with the potential to pay off in a big way, delivering on the promise of gene therapy for disorders currently just out of reach. However, with great promise comes great complexity, and the biotech industry will need to navigate the intricate maze of technical and safety challenges to bring this promising approach to fruition.
Read more from pmc.ncbi.nlm.nih.gov
