The importance of cellular degradation kinetics for understanding mechanisms in targeted protein degradation

Riching, Kristin M.; Caine, Elizabeth A.; Urh, Marjeta; Daniels, Danette L.

doi:10.1039/d2cs00339b

Cited by 25 publications

(17 citation statements)

References 76 publications

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“…One useful aspect of using CC heterodimers in this setting is, as mentioned, that they can be designed to have a desired affinity. The affinity between the substrate and E3 ligase is an important factor in designing synthetic bifunctional degraders (PROTACs) . The use of CC heterodimers enabled us to change the affinity by selecting the appropriate coiled-coil heterodimer.…”

Section: Resultsmentioning

confidence: 99%

Subunits of an E3 Ligase Complex as Degrons for Efficient Degradation of Cytosolic, Nuclear, and Membrane Proteins

Verbič,

Lebar,

Praznik

et al. 2024

ACS Synth. Biol.

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Protein degradation is a highly regulated cellular process crucial to enable the high dynamic range of the response to external and internal stimuli and to balance protein biosynthesis to maintain cell homeostasis. Within mammalian cells, hundreds of E3 ubiquitin ligases target specific protein substrates and could be repurposed for synthetic biology. Here, we present a systematic analysis of the four protein subunits of the multiprotein E3 ligase complex as scaffolds for the designed degrons. While all of them were functional, the fusion of a fragment of Skp1 with the target protein enabled the most effective degradation. Combination with heterodimerizing peptides, protease substrate sites, and chemically inducible dimerizers enabled the regulation of protein degradation. While the investigated subunits of E3 ligases showed variable degradation efficiency of the membrane and cytosolic and nuclear proteins, the bipartite SSD (SOCSbox-Skp1(ΔC111)) degron enabled fast degradation of protein targets in all tested cellular compartments, including the nucleus and plasma membrane, in different cell lines and could be chemically regulated. These subunits could be employed for research as well as for diverse applications, as demonstrated in the regulation of Cas9 and chimeric antigen receptor proteins.

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Section: Resultsmentioning

confidence: 99%

Subunits of an E3 Ligase Complex as Degrons for Efficient Degradation of Cytosolic, Nuclear, and Membrane Proteins

Verbič,

Lebar,

Praznik

et al. 2024

ACS Synth. Biol.

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“…In the first part of her lecture, she focused on cellular kinetic degradation profiles, [21] with an interactive session where members of the course tried to predict profiles based upon differential mechanistic characteristics of degraders [20b] . The discussion focused upon parameters beyond DC 50 (concentration achieving 50 % of total protein degradation) including rates, target resynthesis, native half‐lives, concentration dependency, and hook effect [20b] . She shared lessons learnt from the development of cellular degrader technologies at Promega Corporation for the study of cellular ternary complex, ubiquitination, permeability, and residence time and demonstrated how these enabled understanding of mechanism of action of trivalent PROTACs [22] …”

Section: Presentationsmentioning

confidence: 99%

Section: Presentationsmentioning

confidence: 99%

The 17^th EFMC Short Course on Medicinal Chemistry on Small Molecule Protein Degraders

Ciulli,

O'Connor,

Chung

et al. 2023

ChemMedChem

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The 17th EFMC Short Course on Medicinal Chemistry took place April 23–26, 2023 in Oegstgeest, near Leiden in the Netherlands. It covered for the first time the exciting topic of Targeted Protein Degradation (full title: Small Molecule Protein Degraders: A New Opportunity for Drug Design and Development). The course was oversubscribed, with 35 attendees and 6 instructors mainly from Europe but also from the US and South Africa, and representing both industry and academia. This report summarizes the successful event, key lectures given and topics discussed.

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“…The pVHL30 level begins to recover 12 hours after treatment (Figure S4). Thus, it seems likely that this recovery is due to some type of cell compensation mechanism 49 rather than the instability of the SAPTAC units.…”

Section: Assembly Of Protacs Via Boronate Ester Formationmentioning

confidence: 99%

Self-Assembly of Proteolysis Targeting Chimeras Via Reversible Bioorthogonal Reactions

Gui

Giardina

Balzarini

et al. 2022

Preprint

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Targeted protein degradation (TPD) by PROteolysis TArgeting Chimeras (PROTACs) is of great interest for probe molecule and drug development. However, current bivalent PROTACs are rule-breaking molecules with sub-optimal cellular permeability, solubility, and other drug-like properties. In this study, we report a novel approach for TPD by Self-Assembled Proteolysis TArgeting Chimeras (SAPTACs) in which the target protein and E3 Ubiquitin ligase ligands assemble in cellulo via reversible, bioorthogonal reactions. SAPTACs that me-diate the degradation of the Von Hippel Landau (VHL) E3 Ubiquitin ligase are described. We show that pseu-do-homodimeric degraders for VHL can be assembled in situ through the interaction of VHL ligands linked to phenylboronic acid and catechol as well as to o-acetlyphenylboronic acid and an alkoxyamine. The efficiency of VHL degradation by these SAPTACs is linked to the strength of the reversible covalent interaction.

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The importance of cellular degradation kinetics for understanding mechanisms in targeted protein degradation

Abstract: Targeted protein degradation is a dynamic process regulated not only by the kinetics and mechanisms of the degrader compound, but also the native homeostasis and cellular regulation of the target protein. Image created with BioRender.com.

Cited by 25 publications

References 76 publications

Subunits of an E3 Ligase Complex as Degrons for Efficient Degradation of Cytosolic, Nuclear, and Membrane Proteins

Subunits of an E3 Ligase Complex as Degrons for Efficient Degradation of Cytosolic, Nuclear, and Membrane Proteins

The 17^th EFMC Short Course on Medicinal Chemistry on Small Molecule Protein Degraders

Self-Assembly of Proteolysis Targeting Chimeras Via Reversible Bioorthogonal Reactions

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The importance of cellular degradation kinetics for understanding mechanisms in targeted protein degradation

Abstract: Targeted protein degradation is a dynamic process regulated not only by the kinetics and mechanisms of the degrader compound, but also the native homeostasis and cellular regulation of the target protein. Image created with BioRender.com.

Cited by 25 publications

References 76 publications

Subunits of an E3 Ligase Complex as Degrons for Efficient Degradation of Cytosolic, Nuclear, and Membrane Proteins

Subunits of an E3 Ligase Complex as Degrons for Efficient Degradation of Cytosolic, Nuclear, and Membrane Proteins

The 17th EFMC Short Course on Medicinal Chemistry on Small Molecule Protein Degraders

Self-Assembly of Proteolysis Targeting Chimeras Via Reversible Bioorthogonal Reactions

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Product

Resources

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The 17^th EFMC Short Course on Medicinal Chemistry on Small Molecule Protein Degraders