Three optimized assays for the evaluation of compounds that can rescue p53 mutants

Identifying drugs targeting p53 remains a major focus of precision oncology, with over twenty compounds that can rescue p53 mutants reported. Here, we suggest three easily accessible assays to determine the thermostability, protein folding, and transcriptional activity of p53 mutants—the go-to criteria for evaluating a rescue compound that acts by increasing p53 thermostability. Because of the diversity of p53 mutants, a compound that meets the criteria of one assay does not necessarily meet the criteria of the other assays. For complete details on the use and execution of this protocol, please refer to Chen et al. (2021). [Display omitted] •Optimized and sensitive protocols to evaluate structural mutant p53-rescuing compounds•Choose appropriate p53 mutants in a specific assay to avoid false negatives•Mutant p53 thermostabilizations in DSF assay do not necessarily lead to cellular rescue Identifying drugs targeting p53 remains a major focus of precision oncology, with over twenty compounds that can rescue p53 mutants reported. Here, we suggest three easily accessible assays to determine the thermostability, protein folding, and transcriptional activity of p53 mutants—the go-to criteria for evaluating a rescue compound that acts by increasing p53 thermostability. Because of the diversity of p53 mutants, a compound that meets the criteria of one assay does not necessarily meet the criteria of the other assays.