Development of Photolenalidomide for Cellular Target Identification

Area of Science:

• Chemical Biology
• Proteomics
• Molecular Pharmacology

Background:

• Lenalidomide (Len) is a thalidomide analogue targeting cereblon (CRBN), a key component of the CRL4 E3 ubiquitin ligase complex.
• Understanding the full spectrum of CRBN-interacting proteins is crucial for developing novel therapeutics.

Purpose of the Study:

• To develop a chemical probe for identifying novel targets of lenalidomide and its analogues.
• To investigate the interactions of lenalidomide with its binding partners using chemical proteomics.

Main Methods:

• Development of photolenalidomide (pLen), a photoaffinity probe incorporating a photoactivatable group and an enrichment handle.
• Application of pLen in chemical proteomics experiments using multiple myeloma and HEK293T cell lines.
• Analysis of target engagement and complex formation using molecular modeling and binding site mapping.

Main Results:

• Photolenalidomide (pLen) retains the biological activity and CRBN-binding properties of lenalidomide (Len).
• pLen successfully identified known targets (IKZF1, CRBN) and a novel target, eukaryotic translation initiation factor 3 subunit i (eIF3i).
• eIF3i forms a ternary complex with CRBN upon lenalidomide treatment but is not ubiquitylated or degraded.

Conclusions:

• Photolenalidomide (pLen) is an effective chemical probe for identifying both known and novel CRBN-interacting proteins.
• Ligand binding to CRBN can induce interactions with proteins that are not necessarily degraded, broadening the scope of target identification.
• This approach has broad translatability for discovering new therapeutic targets in various biological systems.