(T1430-10-63) Impact of Preparation Technique and Storage Condition on the Quality of Suspension Vehicles Used in Preclinical In Vivo Studies

Purpose: Preclinical research facilities responsible for in vivo PK/PD or toxicity studies are frequently faced with a high throughput of samples and tight timelines. To safe time, vehicles are oftentimes prepared and stored under various conditions already days before the actual application. While the chemical stability of the API in the vehicle is usually known, changes that affect physical integrity in a suspension formulation, such as homogeneity and particle growth, are checked, if at all, only macroscopically. However, unidentified changes can have devastating effects on reproducibility as well as drug dissolution and, ultimately, on the resulting bioavailability. The aim of this research was to gain insights into the impact of preparation methods and storage conditions in two commonly used vehicles for animal studies.

Methods: Suspension formulations of two different drug substances in two different vehicles consisting of 0.5% Methocel and 0.25% Tween 20 in PBS (vehicle 1) and 0.25% Soluplus in PBS (vehicle 2) were prepared with a concentration of 10 mg/mL, using different preparation techniques (i.e., fast homogenization via UltraTurrax® or simple stirring with a magnetic stirring bar) and subsequently stored under various conditions for 7 days. These formulations were stored at two different storage conditions, stirring at room temperature and storage in the fridge without stirring. Two representative model compounds were used, one of which being a neutral API (Compound A) with a logP of 3.5 and the other one being a slightly acidic API (Compound B) with a logP of 1.1. Changes in suspension were investigated using macroscopic and microscopic imaging.

Results: Both compounds showed macroscopic sedimentation to various degrees over the course of 7 days for both preparation techniques. While the stirred samples showed only marginal sedimentation, unstirred conditions at 8 °C led to pronounced sedimentation. After redispersion by vortexing, however, homogeneous white suspensions could be restored for all samples, according to macroscopic examination. Microscopic imaging for compound A revealed particle growth over a period of 72 h for vehicles prepared by stirring, with a more pronounced growth visible for storage at 8 °C. Using the UltraTurrax® device for formulation preparation, particle growth over time was only detected in vehicle 2 at a storage temperature of 8 °C. Compound B demonstrated no substantial difference in preparation method or storage condition over 72 h in microscopic imaging. After 7 days, a slight particle growth in vehicle 2 was observed in samples of both preparation methods and all storage conditions, with the exception being stirred at room temperature after preparation using the UltraTurrax®.

Conclusion: While both vehicles in combination with the model APIs did not show relevant macroscopic changes in appearance, microscopic imaging revealed that both preparation techniques and storage conditions can substantially alter particle properties over time, e.g. particle growth by Ostwald ripening, and therefore endanger reproducibility and in vivo performance. As a cheap and fast mitigation strategy, research facilities should routinely monitor particle behavior of compounds in suspension over a relevant storage time by microscopic imaging. Depending on the compound, the obtained results suggest that the preparation of suspensions by means of the UltraTurrax® as well as storing the suspension formulations at room temperature may be clearly preferred over the preparation by stirring and storing in the fridge. Future research on this topic should also consider the impact of suspension formulation preparation methods and storage conditions on further parameters such as solid-state properties, such as possible polymorph changes or conversion of crystalline to amorphous state which may also have a substantial effect on the in vivo performance.