Recombinant adeno-associated virus (rAAV) has emerged as the leading gene delivery platform for treatment of monogenic disorders. Currently, for clinical and commercial products, rAAVs are typically formulated and stored below -65 °C as frozen liquid. Their long-term storage is often far from ideal because it may result in shorter drug product (DP) shelf-life compared to recombinant protein-based biologics, and also presents challenges for supply chain and inventory management. Consequently, there is great interest in developing robust lyophilized AAV DPs that are stable at 2 to 8 °C. In this study, we evaluated formulation excipients required for stable lyophilized AAV8 products including buffers, salts, cryoprotectants/lyoprotectants, surfactants, and bulking agents, and optimized the concentrations and ratios between the excipients. This led to the identification of the lead formulation that demonstrated short-term in-solution stability at 25 °C and, upon lyophilization, sufficient long-term stability at 2 to 8 °C. Our study demonstrated that, in the presence of 110 mM salts, mannitol can serve as an effective bulking agent with the appropriate formulation and lyophilization process design, and the sucrose to mannitol ratio is critical to maintain the stability and cake appearance of the lyophilized AAV8 DP. Thorough characterization of the effect of formulation components on the properties and quality of the lyophilized DP led to an optimized AAV8 lyophilized DP. This approach could be applied to streamline the future development of lyophilized AAV gene therapy products with various target transgenes and capsid serotypes.
Keywords: Adeno-associated virus (AAV); Formulation excipient; Freeze-drying; Lyophilization; Mannitol; Vector genome (vg) titer.
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