Intracellular cyclization-coupled peptide library screening yields potent transcription factor antagonists.

Area of Science:

• Drug discovery and development
• Molecular biology
• Chemical biology

Background:

• Transcription factors possess flat, dynamic interfaces, making them challenging targets for drug discovery.
• Developing effective therapeutics against such targets requires novel strategies for generating constrained peptides.

Purpose of the Study:

• To present an intracellular cyclization strategy for in-cell generation of conformationally constrained peptide libraries.
• To develop a platform for simultaneous selection of peptide sequence and constraint site, eliminating iterative synthesis.
• To discover peptide antagonists for the oncogenic transcription factor CREB1.

Main Methods:

• Utilizing bis-alkylating reagents for post-translational peptide stapling within bacterial membranes.
• Integrating intracellular cyclization with the transcription block survival (TBS) assay to create the icTBS platform.
• Screening peptide libraries against CREB1 to identify nanomolar-affinity antagonists.

Main Results:

• The icTBS platform successfully generated conformationally constrained peptide libraries in vivo.
• Three nanomolar-affinity CREB1 antagonists were identified, with cyclized variants showing enhanced cellular activity.
• The lead peptide demonstrated anti-cancer effects, including suppressed CREB1 activity and induced apoptosis in cancer cells.

Conclusions:

• Intracellular cyclization provides a general, genetically encoded route to discover constrained peptide therapeutics.
• The icTBS platform enables the targeting of previously undruggable protein-DNA interfaces.
• This strategy holds promise for developing novel peptide-based cancer therapies.