A High-Throughput Small Molecule Screen Identifies Targets That Increase AAV9 Production In Suspension HEK293 Cells

By Francis Grafton, Ryan Feitzinger, Kaylin Fisher, Lily Leveque-Eichhorn, Christopher A. Reid, and Mohammad A. Mandegar

Recombinant AAV is widely used as a vector for gene delivery, with over 900 pre-clinical and clinical programs in progress; however, inefficient manufacturing methods contribute to high costs, limiting the availability of gene therapies. In this study, we outline a high-throughput small molecule screening strategy to identify compounds that enhance AAV9 production in cells.

Using the ATLAS (Arrayed Targeted Library for AAV Screening) platform, we screened over 3,000 small molecules and identified key targets, including transmembrane proteins, DNA repair proteins, cell-cycle regulators, and epigenetic modulators. The top 71 compounds were further assessed for dose-response effects in our proprietary clonal HEK293 cell line (AC001.230). Among these, we discovered a novel compound, SM-016, which significantly increased rAAV9 production in a dose-dependent manner. Ongoing studies will explore SM-016's effects on different AAV serotypes and its potential for large-scale manufacturing, while also analyzing its impact on the quality attributes of AAV vector batches.