For AAV programs, our pre-development testing has typical been very minimal – this is causing substantial developability challenges for our programs and wasted people-power to resolve issues that could be avoided. An understanding of where degradation transitions occur in {concentration, temperature, pH, ionic strength} phase space is vital before development begins. For example, a lack of understand of low ionic strength tolerance may cause method-induced artifacts in a PCR titer method that dilutes into a low-salt buffer before measurement. This talk introduces high-throughput, microplate-based methods to aid in the measurement of degradation transitions based on intrinsic and extrinsic fluorescence in addition to anisothermal dynamic light scattering and shows how they have been used to solve developability challenges and increase our product understanding. These methods could be easily implemented in pre-development testing or in your group!
A high degree of dependence was found for serotype and for formulation conditions such as concentration and ionic strength in the position of the degradation transitions with respect to almost all stresses tested. As concentration and buffer conditions are frequently altered in the sample manipulation required by most analytical methods, the results reinforce the important of understanding where degradation transitions occur. The serotype dependence suggest that AAV development may be less amenable to platform approaches. The high risk of method-induced artifacts necessitates the addition of first-principles methods to our analytical panels – for example we cannot rely solely on PCR for titer measurements. Furthermore, for every serotype tested, the genome ejection transition preceded all other transitions - Sanofi and the industry in general could benefit from applying more concern to genome ejection and less concern to aggregation in the development of analytical panels.