Objective

Comprehensive analyses of complex cellular signaling cascades are essential to get a mechanistic understanding of the processes underlying drug response and drug resistance. However, there are few technologies available for such in-depth pathway analyses on the level of phospho proteins, and therefore, profiling efforts are all too often focused on genomics methods.

Using 20-60 microgram of total protein per cell or tissue sample, the DigiWest^® immuno-assay technology (Treindl et al., Nature Comm 2016; www.digiwest.de) enables parallelized analyses of currently up to 800 total and phospho proteins per sample, through a combination of Western blotting with the multiplexing ability of the Luminex^® platform, thus allowing for comprehensive pathway profiling data beyond genomics.

We will present sample data from preclinical oncology studies where DigiWest^® has been successfully applied for lead compound characterization and biomarker discovery.

Furthermore, to facilitate a wider use of DigiWest^®, we have meanwhile also profiled samples from rat, dog, minipig and cynomolgus origin with a large collection of commercially available antibodies, and we have created pathway panels with hundreds of pre-validated antibodies for each of the six species.

These results represent a basis for improved in vivo pharmacology and preclinical safety assessments, by making investigations in model species more amenable to protein profiling, thereby also contributing to the 3R aims by providing a technology that helps getting the most out of animal experiments.