title: Mechanistic validation of a clinical lead stapled peptide that reactivates p53 by dual HDM2 and HDMX targeting.
abstract: Hydrocarbon-stapled peptides that display key residues of the p53 transactivation domain have emerged as bona fide clinical candidates for reactivating the tumor suppression function of p53 in cancer by dual targeting of the negative regulators HDM2 and HDMX. A recent study questioned the mechanistic specificity of such stapled peptides based on interrogating their capacity to disrupt p53/HDM2 and p53/HDMX complexes in living cells using a new recombinase enhanced bimolecular luciferase complementation platform (ReBiL). Here, we directly evaluate the cellular uptake, intracellular targeting selectivity and p53-dependent cytotoxicity of the clinical prototype ATSP-7041. We find that under standard serum-containing tissue culture conditions, ATSP-7041 achieves intracellular access without membrane disruption, dose-dependently dissociates both p53/HDM2 and p53/HDMX complexes but not an unrelated protein complex in long-term ReBiL experiments, and is selectively cytotoxic to cancer cells bearing wild-type p53 by inducing a surge in p53 protein level. These studies underscore the importance of a thorough stepwise approach, including consideration of the time-dependence of cellular uptake and intracellular distribution, in evaluating and advancing stapled peptides for clinical translation.
authors.affiliation: Department of Pediatric Oncology and the Linde Program in Cancer Chemical Biology, Dana-Farber Cancer Institute, Boston, MA, USA.
authors.forename: F
authors.initials: F
authors.lastname: Wachter
authors.affiliation: Department of Pediatric Oncology and the Linde Program in Cancer Chemical Biology, Dana-Farber Cancer Institute, Boston, MA, USA.
authors.forename: A M
authors.initials: AM
authors.lastname: Morgan
authors.affiliation: Department of Pediatric Oncology and the Linde Program in Cancer Chemical Biology, Dana-Farber Cancer Institute, Boston, MA, USA.
authors.forename: M
authors.initials: M
authors.lastname: Godes
authors.affiliation: Department of Pediatric Oncology and the Linde Program in Cancer Chemical Biology, Dana-Farber Cancer Institute, Boston, MA, USA.|The Harvard-MIT Program in Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA, USA.
authors.forename: R
authors.initials: R
authors.lastname: Mourtada
authors.affiliation: Department of Pediatric Oncology and the Linde Program in Cancer Chemical Biology, Dana-Farber Cancer Institute, Boston, MA, USA.
authors.forename: G H
authors.initials: GH
authors.lastname: Bird
authors.affiliation: Department of Pediatric Oncology and the Linde Program in Cancer Chemical Biology, Dana-Farber Cancer Institute, Boston, MA, USA.
authors.forename: L D
authors.initials: LD
authors.lastname: Walensky
mesh_terms: D000970:Antineoplastic Agents / Q000493:pharmacokinetics / Q000494:pharmacology; D001859:Bone Neoplasms / Q000188:drug therapy / Q000378:metabolism; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism*; D012516:Osteosarcoma / Q000188:drug therapy / Q000378:metabolism; D010456:Peptides, Cyclic / Q000493:pharmacokinetics / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
chemical_list: C584947:ATSP-7041; D000970:Antineoplastic Agents; D018797:Cell Cycle Proteins; C482775:MDM4 protein, human; D009687:Nuclear Proteins; D010456:Peptides, Cyclic; D011518:Proto-Oncogene Proteins; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2