Don't shake it! Mechanical stress testing of mRNA-lipid nanoparticles.

ABSTRACT

Shaking stress studies are typically performed during formulation development to test the liability of a drug product towards interfacial stress occurring during transport, especially if a liquid formulation is desired. We evaluated various shaking procedures using a polyA-surrogate solution and verified our findings by eGFP-LNP cell-expression experiments. Shaking on an orbital shaker in vertical and horizontal orientations at increasing speeds from 300 to 600 rpm resulted in decreasing levels of encapsulated nucleic acid content, larger LNP sizes, and decreasing PDI. We report that vertical and horizontal shaking of both polyA- and eGFP-LNPs led to white deposits on the inner glass vial surface, depending on time, rpm, and temperature. Increasing the fill volume/smaller headspace (0.3 versus 0.9 mL fill) did not mitigate this phenomenon in the studied configuration, and the use of hydrophobic primary packaging even accelerated the formation of white deposits. In contrast, we demonstrated that a lyophilized polyA-LNP dosage form was less susceptible to shaking and maintained cake integrity and product properties. Multiple vortexing steps resulted in an increase in LNP size, PDI, and a decrease in encapsulated polyA content. We conclude that shaking experiments of nucleic acid-loaded LNPs in their final configuration at intended transport conditions need to be considered during technical development.