title: Catalytic in vivo protein knockdown by small-molecule PROTACs.
abstract: The current predominant therapeutic paradigm is based on maximizing drug-receptor occupancy to achieve clinical benefit. This strategy, however, generally requires excessive drug concentrations to ensure sufficient occupancy, often leading to adverse side effects. Here, we describe major improvements to the proteolysis targeting chimeras (PROTACs) method, a chemical knockdown strategy in which a heterobifunctional molecule recruits a specific protein target to an E3 ubiquitin ligase, resulting in the target's ubiquitination and degradation. These compounds behave catalytically in their ability to induce the ubiquitination of super-stoichiometric quantities of proteins, providing efficacy that is not limited by equilibrium occupancy. We present two PROTACs that are capable of specifically reducing protein levels by >90% at nanomolar concentrations. In addition, mouse studies indicate that they provide broad tissue distribution and knockdown of the targeted protein in tumor xenografts. Together, these data demonstrate a protein knockdown system combining many of the favorable properties of small-molecule agents with the potent protein knockdown of RNAi and CRISPR.
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA.
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Bondeson
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Alina
authors.initials: A
authors.lastname: Mares
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Ian E D
authors.initials: IE
authors.lastname: Smith
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA.
authors.forename: Eunhwa
authors.initials: E
authors.lastname: Ko
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Sebastien
authors.initials: S
authors.lastname: Campos
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Afjal H
authors.initials: AH
authors.lastname: Miah
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Katie E
authors.initials: KE
authors.lastname: Mulholland
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Natasha
authors.initials: N
authors.lastname: Routly
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA.
authors.forename: Dennis L
authors.initials: DL
authors.lastname: Buckley
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA.
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Gustafson
authors.affiliation: Cellzome, a GSK company, Heidelberg, Germany.
authors.forename: Nico
authors.initials: N
authors.lastname: Zinn
authors.affiliation: Cellzome, a GSK company, Heidelberg, Germany.
authors.forename: Paola
authors.initials: P
authors.lastname: Grandi
authors.affiliation: Cellzome, a GSK company, Heidelberg, Germany.
authors.forename: Satoko
authors.initials: S
authors.lastname: Shimamura
authors.affiliation: Cellzome, a GSK company, Heidelberg, Germany.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Bergamini
authors.affiliation: Cellzome, a GSK company, Heidelberg, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Faelth-Savitski
authors.affiliation: Cellzome, a GSK company, Heidelberg, Germany.
authors.forename: Marcus
authors.initials: M
authors.lastname: Bantscheff
authors.affiliation: Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA.
authors.forename: Carly
authors.initials: C
authors.lastname: Cox
authors.affiliation: Arvinas, Inc., New Haven, Connecticut, USA.
authors.forename: Deborah A
authors.initials: DA
authors.lastname: Gordon
authors.affiliation: Arvinas, Inc., New Haven, Connecticut, USA.
authors.forename: Ryan R
authors.initials: RR
authors.lastname: Willard
authors.affiliation: Arvinas, Inc., New Haven, Connecticut, USA.
authors.forename: John J
authors.initials: JJ
authors.lastname: Flanagan
authors.affiliation: Pattern Recognition Receptor Discovery Performance Unit, GlaxoSmithKline, Collegeville, Pennsylvania, USA.
authors.forename: Linda N
authors.initials: LN
authors.lastname: Casillas
authors.affiliation: Pattern Recognition Receptor Discovery Performance Unit, GlaxoSmithKline, Collegeville, Pennsylvania, USA.
authors.forename: Bartholomew J
authors.initials: BJ
authors.lastname: Votta
authors.affiliation: Division of Biology &Biological Engineering, California Institute of Technology, Pasadena, California, USA.
authors.forename: Willem
authors.initials: W
authors.lastname: den Besten
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Kristoffer
authors.initials: K
authors.lastname: Famm
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Laurens
authors.initials: L
authors.lastname: Kruidenier
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Paul S
authors.initials: PS
authors.lastname: Carter
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: John D
authors.initials: JD
authors.lastname: Harling
authors.affiliation: GSK Medicines Research Centre, Stevenage, UK.
authors.forename: Ian
authors.initials: I
authors.lastname: Churcher
authors.affiliation: 1] Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut, USA. [2] Department of Chemistry, Yale University, New Haven, Connecticut, USA. [3] Department of Pharmacology, Yale University, New Haven, Connecticut, USA.
authors.forename: Craig M
authors.identifier: 0000000284562005
authors.initials: CM
authors.lastname: Crews
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001665:Binding Sites; D055162:Biocatalysis; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D005260:Female; D006801:Humans; D061986:MCF-7 Cells; D051379:Mice; D008958:Models, Molecular; D058990:Molecular Targeted Therapy; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D009368:Neoplasm Transplantation; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D059748:Proteolysis; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D011960:Receptors, Estrogen / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D054852:Small Molecule Libraries / Q000494:pharmacology*; D025801:Ubiquitin / Q000235:genetics / Q000378:metabolism; D054875:Ubiquitination; D051794:Von Hippel-Lindau Tumor Suppressor Protein / Q000235:genetics / Q000378:metabolism; D000097983:ERRalpha Estrogen-Related Receptor
chemical_list: D000970:Antineoplastic Agents; D009363:Neoplasm Proteins; D046988:Proteasome Endopeptidase Complex; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D011960:Receptors, Estrogen; D054852:Small Molecule Libraries; D025801:Ubiquitin; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D000097983:ERRalpha Estrogen-Related Receptor; C506253:RIPK2 protein, human; C497446:VHL protein, human