A Chemoproteomic Strategy for Direct and Proteome-Wide Covalent Inhibitor Target-Site Identification.

Autor:	Browne CM; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States., Jiang B; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States., Ficarro SB; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States.; Blais Proteomics Center , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States., Doctor ZM; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States., Johnson JL; Meyer Cancer Center , Weill Cornell Medicine and New York Presbyterian Hospital , New York , New York 10065 , United States., Card JD; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Blais Proteomics Center , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States., Sivakumaren SC; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States., Alexander WM; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Blais Proteomics Center , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States., Yaron TM; Meyer Cancer Center , Weill Cornell Medicine and New York Presbyterian Hospital , New York , New York 10065 , United States., Murphy CJ; Meyer Cancer Center , Weill Cornell Medicine and New York Presbyterian Hospital , New York , New York 10065 , United States., Kwiatkowski NP; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States.; Whitehead Institute for Biomedical Research , Cambridge , Massachusetts 02142 , United States., Zhang T; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States., Cantley LC; Meyer Cancer Center , Weill Cornell Medicine and New York Presbyterian Hospital , New York , New York 10065 , United States., Gray NS; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Biological Chemistry and Molecular Pharmacology , Harvard Medical School , Boston , Massachusetts 02115 , United States., Marto JA; Department of Cancer Biology , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Blais Proteomics Center , Dana-Farber Cancer Institute , Boston , Massachusetts 02215 , United States.; Department of Pathology , Brigham and Women's Hospital, Harvard Medical School , Boston , Massachusetts 02115 , United States.
Jazyk:	angličtina
Zdroj:	Journal of the American Chemical Society [J Am Chem Soc] 2019 Jan 09; Vol. 141 (1), pp. 191-203. Date of Electronic Publication: 2018 Dec 20.
DOI:	10.1021/jacs.8b07911
Abstrakt:	Despite recent clinical successes for irreversible drugs, potential toxicities mediated by unpredictable modification of off-target cysteines represents a major hurdle for expansion of covalent drug programs. Understanding the proteome-wide binding profile of covalent inhibitors can significantly accelerate their development; however, current mass spectrometry strategies typically do not provide a direct, amino acid level readout of covalent activity for complex, selective inhibitors. Here we report the development of CITe-Id, a novel chemoproteomic approach that employs covalent pharmacologic inhibitors as enrichment reagents in combination with an optimized proteomic platform to directly quantify dose-dependent binding at cysteine-thiols across the proteome. CITe-Id analysis of our irreversible CDK inhibitor THZ1 identified dose-dependent covalent modification of several unexpected kinases, including a previously unannotated cysteine (C840) on the understudied kinase PKN3. These data streamlined our development of JZ128 as a new selective covalent inhibitor of PKN3. Using JZ128 as a probe compound, we identified novel potential PKN3 substrates, thus offering an initial molecular view of PKN3 cellular activity. CITe-Id provides a powerful complement to current chemoproteomic platforms to characterize the selectivity of covalent inhibitors, identify new, pharmacologically addressable cysteine-thiols, and inform structure-based drug design programs.
Databáze:	MEDLINE
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