- Project team: Yohei Yokobayashi, Elvira Vitu, Haifeng Zhang
- Host unit: Nucleic Acid Chemistry and Engineering Unit
- Project duration: 1 April 2026 – 31 March 2027
- Keywords: Gene therapy, AAV, Riboswitch, Aptamer
- Previous project stage: Optimization of AAV Vector Production Using Riboswitch (Seed Phase)
Riboswitch-Mediated Regulation of AAV Rep and Cap Expression to Improve Viral Vector Manufacturing
Problem
Adeno-associated virus (AAV) vectors are emerging as a major class of gene therapy vectors used to deliver therapeutic genes to the patient’s cells. However, manufacturing of AAV vectors is complex and expensive. One of the major challenges in AAV manufacturing is the high level of “empty capsids” or virus shells containing no genes (DNA) during production (Figure 1).
These empty capsids necessitate costly purification processes and contribute to increased inflammation upon administration. Therefore, solutions to minimize empty capsids while improving full capsid yield will benefit both AAV manufacturers and patients.
Figure 1. Full capsids contain single-stranded DNA that encodes therapeutic genes while empty capsids do not.
Solution
We aim to address the empty capsid problem by controlling the expression level and timing of viral proteins using “riboswitches.” Riboswitches are gene switches that can induce or suppress protein expression in cultured cells by the addition of a small molecule.
Our riboswitch technology is based on the new small molecule – RNA aptamer pairs that were discovered in our group (Figure 2), which exhibit excellent switch characteristics. We believe that by precisely manipulating the expression of various viral genes during AAV production in cultured cells, we can improve vector production efficiency and quality.
Figure 2. Small molecule (ASP2905) and the RNA aptamer (AC17-4) that are used to construct riboswitches that regulate gene expression.