Development and Application of an Automated Solution Stability Assay for Drug Discovery

Abstract

Screening of solution stability provides an early alert on potential liabilities of drug candidates so that strategies can be developed to overcome the challenges. A fully automated solution stability assay has been developed to accelerate traditional manual operation. The assay uses the advanced capabilities of a high-performance liquid chromatography instrument that is present in many pharmaceutical research laboratories. The samples are prepared automatically by a temperature-controlled autosampler. The samples are delivered to the stability matrices, mixed, incubated, and injected at selected time points during the reaction time course. This automated process occurs without operator intervention, thus allowing 96 experiments to be run with 0.5 h of a scientist's time compared to 8 h for the same study when performed manually. Automation not only eliminates the manual operation but also improves accuracy and throughput. The assay protocol has been optimized to achieve homogenous mixing and eliminate carryover. The assay is robust, flexible, and high throughput. It can be used to study stability for a large number of samples under multiple incubation conditions and has a wide range of applications in drug discovery and development, such as screening compound stability in biological assay media, obtaining a stability-pH profile, surveying compound stability in physiological fluids, and performing development forced degradation and excipient compatibility. (Journal of Biomolecular Screening 2006:40-47)

Solution Stability Methods

“…5 Into the autosampler are placed two 96-well plates, one containing dimethylsulfoxide (DMSO) stock solutions of the test compounds and the other containing wells filled (e.g., 180 μL) with the stability solution ( Figure 31.1). 5 Into the autosampler are placed two 96-well plates, one containing dimethylsulfoxide (DMSO) stock solutions of the test compounds and the other containing wells filled (e.g., 180 μL) with the stability solution ( Figure 31.1).…”

High Throughput Screening in the Twenty-First Century

“…One factor in the current uncertainty about storage conditions is the significant cost associated with maintaining an effective quality control program to monitor the purity and integrity of screening libraries ranging from half a million to a million and a half compounds. Despite reports of automated analytical systems, the time and expense it takes to determine the status of every compound on a rotating schedule is more than most organizations are willing to spend [57][58][59]. Ultimately, as low volume liquid handling instrumentation has improved and the miniaturization of precision labware has evolved, the preparation of multiple replicate copies of small aliquots of the screening library in plates and microtubes designed for one-time use may obviate some concerns surrounding storage conditions and freeze/thaw cycles.…”

Utility of Mass Spectrometry for Pharmaceutical Profiling Applications

“…This allows studies of solution stability for 24 compounds or 24 buffer conditions. Di, et al [46] developed an on-line sample preparation system using an HPLC autosampler for sample delivery, mixing, incubation and injection. The method was developed using HPLC systems and software that are present in most pharmaceutical R & D laboratories.…”

“…High sensitivity for methods requiring low LOQ Specificity Increased specificity by detection of the molecular ion or MS/MS product ion, so that impurities do not cause erroneous results [14] Specificity Individual analysis of components in combinatorial mix (cassette) [12,14,16,22] Specificity Allows use of DMSO, excipients, proteins and other matrix components that would interfere with detection by UV [13,32] Specificity Use of internal standards for quantitation or retention time standardization [12,20] Specificity Obtaining initial structural data (e.g., MW) on test compounds and unknown degradants produced, in addition to quantitating the test compound [32] Cost Easy integration with HPLC autosamplers, valves and columns for on-line sample incubation, physicochemical experiments, purity quantitation and cleanup [12,16,20,22,23,45,46,52] widely varying response to diverse components. ELSD responses were more universal than UV above MW 300, however, many synthetic intermediates or starting materials have MW below 300.…”