Improved phosphoproteomic analysis for phosphosignaling and active-kinome profiling in Matrigel-embedded spheroids and patient-derived organoids.

ABSTRACT: Many attempts have been made to reproduce the three-dimensional (3D) cancer behavior. For that purpose, Matrigel, an extracellular matrix from Engelbreth-Holm-Swarm mouse sarcoma cell, is widely used in 3D cancer models such as scaffold-based spheroids and patient-derived organoids. However, severe ion suppression caused by contaminants from Matrigel hampers large-scale phosphoproteomics. In the present study, we successfully performed global phosphoproteomics from Matrigel-embedded spheroids and organoids. Using acetone precipitations of tryptic peptides, we identified more than 20,000 class 1 phosphosites from HCT116 spheroids. Bioinformatic analysis revealed that phosphoproteomic status are significantly affected by the method used for the recovery from the Matrigel, i.e., Dispase or Ce

SUBMITTER: Abe Y 

PROVIDER: S-EPMC6065387 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature