Dataset Information

Identification of an allosteric site on the E3 ligase adapter cereblon

ABSTRACT: Cereblon (CRBN) is the target of thalidomide derivatives that achieve therapeutic efficacy against some hematologic neoplasias by recruiting neosubstrates for degradation. Despite the intense investigation of orthosteric thalidomide derivatives, little is known about alternate binding sites on CRBN. Here, we report the discovery of an evolutionarily conserved cryptic allosteric binding site on CRBN. Small molecule SB-405483 binds the allosteric site to cooperatively enhance the binding of orthosteric ligands and alter their neosubstrate degradation profiles. A survey of over 100 orthosteric ligands and their degradation targets reveals trends in the classes of compounds and neosubstrates where degradation outcomes are enhanced or inhibited by SB-405483. Structural investigations provide a mechanistic basis for the effects of the allosteric ligand by shifting the conformational distribution of CRBNopen to a novel CRBNint and increasing the CRBNclosed state. The discovery of a cryptic allosteric binding site on CRBN that alters the functional effects of orthosteric ligands opens new directions with broad implications for improving the selectivity and efficacy of CRBN therapeutics.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Christina Woo

LAB HEAD: Christina Woo

PROVIDER: PXD052094 | Pride | 2025-11-10

REPOSITORIES: Pride

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			Action	DRS
	230519E_Vanessa_TMT_MS2_SAM11647.raw	Raw
	230519E_Vanessa_TMT_MS2_SAM11648.raw	Raw
	230519E_Vanessa_TMT_MS2_SAM11649.raw	Raw
	230519E_Vanessa_TMT_MS2_SAM11650.raw	Raw
	230519E_Vanessa_TMT_MS2_SAM11651.raw	Raw

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Publications

Acylation modification-mediated metabolism reprogramming in cancer.

Huang Kaiwei K Wang Yunshan Y Li Bohao B Xiu Yuchen Y Zhan Hanxiang H Wei Guangwei G Duan Yangmiao Y

iScience 20251030 12

With the development of high-resolution liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS), various modifications resulting from the reactivity of acyl-CoA intermediates with protein residues have been identified. Acylation modifications, a diverse type of post-translational modifications (PTMs), play pivotal roles in regulating protein functions and are critically involved in tumor metabolic reprogramming. These modifications exhibit significant regulatory effects in various ...[more]

PMID: 41321623

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Project description:Targeted protein degradation is the selective removal of a protein of interest through hijacking intracellular protein cleanup machinery. This rapidly growing field currently relies heavily on the use of the E3 ligase Cereblon (CRBN) to target proteins for degradation, including the immunomodulatory drugs (IMiDs) thalidomide, lenalidomide, and pomalidomide which work through a molecular glue mechanism of action with CRBN. While CRBN recruitment can result in degradation of a specific protein of interest (e.g. efficacy), degradation of other proteins (called CRBN neosubstrates) also occurs. Degradation of one or more of these CRBN neosubstrates is believed to play an important role in thalidomide-related developmental toxicity observed in rabbits and primates. We identified a set of 25 proteins of interest associated with CRBN-related protein homeostasis and/or embryo/fetal development. We developed a targeted assay for these proteins combining peptide immunoaffinity enrichment and high-resolution mass spectrometry and successfully applied this assay to rabbit embryo samples from pregnant rabbits dosed with three IMiDs. We confirmed previously reported in vivo decreases in neosubstrates like SALL4, as well as provided evidence of neosubstrate changes for proteins only examined in vitro previously. While there were many proteins that were similarly decreased by all three IMiDs, no compound had the exact same neosubstrate degradation profile as another. We compared our data to previous literature reports of IMiD induced degradation and known developmental biology associations. Based on our observations, we recommend monitoring at least a major subset of these neosubstrates in a developmental test system to improve CRBN-binding compound-specific risk assessment. A strength of our assay is that it is configurable, and the target list can be readily adapted to focus on only a subset of proteins of interest or expanded to incorporate new findings as additional information about CRBN biology is discovered.