Fluorescence-Coupled Ubiquitination Assay as a High-Throughput Screening Strategy for Novel Cereblon Degraders
Cereblon (CRBN)-based protein degradation, via molecular glue degraders (MGDs) and proteolysis-targeting chimeras (PROTACs), is a promising cancer treatment strategy in targeted protein degradation (TPD). However, novel degraders discovery remains limited due to the lack of robust, high-throughput screening (HTS) methods for processing pools of purified compounds or complex chemical synthesis mixtures. Here, we introduce an innovative HTS strategy that employs a highly sensitive, fluorescence-coupled ubiquitination assay to identify CRBN-based degraders. This approach tracks ubiquitinated target proteins via gel-based analyses, and thereby progressively narrows down the list of potential degrader molecules from large-scale compound libraries or chemical reaction mixtures. Using this strategy, we identified LL-BPTF-8, a promising lead compound of PROTAC degrader with high potency and selectivity that targets the bromodomain PHD finger transcription factor (BPTF). Overall, our method offers a low-cost, rapid, and versatile platform for the HTS of protein degrader candidates, significantly streamlining the discovery of novel degraders.
Authors
- Hua Lin (ORCID: https://orcid.org/0000-0002-0840-6553)
- Zemin Zhang (ORCID: https://orcid.org/0009-0005-3119-7509)
- Ying Xu (ORCID: https://orcid.org/0009-0004-6539-4431)
- Cheng Luo (ORCID: https://orcid.org/0000-0003-3864-8382)
- Yanan Deng (ORCID: https://orcid.org/0000-0002-1383-4368)
- Ke Xu (ORCID: https://orcid.org/0000-0002-9969-3681)
- Zizhong Zhang (ORCID: https://orcid.org/0000-0002-9541-8981)
- Kun Yang (ORCID: https://orcid.org/0000-0002-6153-7683)
- Shiling Yang
- Huan Xiong (ORCID: https://orcid.org/0009-0002-8920-9355)
- Shijie Chen (ORCID: https://orcid.org/0000-0001-9724-7453)
- Jin Lin (ORCID: https://orcid.org/0000-0003-1595-9161)
- Xiangdong Ding
- Yiran Hu (ORCID: https://orcid.org/0000-0002-2310-4676)
- Zhihai Li (ORCID: https://orcid.org/0000-0002-2816-6243)
- Hesong Xu
- Zhengzheng Ye
- Yan Chen (ORCID: https://orcid.org/0009-0002-3270-358X)
Institutions
- Fujian Normal University (CN)
- Fujian Medical University (CN)
- Guiyang Medical University (CN)
- Chinese Academy of Sciences (CN)
- Shanghai Institute of Materia Medica (CN)
- Drug Discovery Laboratory (Norway) (NO)
- Institute for Advanced Study (DE)
- University of Chinese Academy of Sciences (CN)
- East China Normal University (CN)
Publication Details
- Journal
- Journal of Medicinal Chemistry
- Published
- 2025-05-07
- DOI
- https://doi.org/10.1021/acs.jmedchem.5c00065
- Citations
- 3
- Primary Topic
- Protein Degradation and Inhibitors
- Type
- article
- Field-Weighted Citation Impact
- 2.42
Funders
- National Natural Science Foundation of China
- China Postdoctoral Science Foundation
- Guangdong Science and Technology Department
- National Key Research and Development Program of China
- Natural Science Foundation of Zhejiang Province