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Discovery and Optimization of Small Molecule Inhibitors of the SLIT2/ROBO1 Protein-Protein Interaction Using
Biorxiv : the Preprint Server for Biology
|May 13, 2026
Summary
Researchers developed small molecule inhibitors for the challenging SLIT2-ROBO1 protein-protein interaction using DNA-encoded library screening. Optimization yielded potent modulators targeting cell migration and tumor progression pathways.
Area of Science:
- Biochemistry
- Molecular Biology
- Drug Discovery
Background:
- Extracellular protein-protein interactions (PPIs) are difficult small molecule targets due to large, dynamic interfaces.
- The SLIT2-ROBO1 interaction is crucial for cell migration and tumor progression but remains under-explored for therapeutic intervention.
Purpose of the Study:
- To discover and optimize small molecule inhibitors targeting the SLIT2-ROBO1 interaction.
- To explore the potential of DNA-encoded library (DEL) screening for extracellular PPIs.
Main Methods:
- DNA-encoded library (DEL) screening against SLIT2.
- Affinity selection, orthogonal biophysical assays, and time-resolved FRET assays for validation.
- Molecular dynamics simulations, induced-fit docking, and fragment-based validation for optimization.
Main Results:
- Identified four structurally diverse hit compounds from DEL screening.
- Optimized analog showed a ~50-fold increase in binding affinity and ~9-fold enhancement in functional potency.
- Discovered a minimal azaindole pharmacophore with nanomolar binding affinity.
Conclusions:
- DEL screening is effective for discovering inhibitors of challenging extracellular PPIs.
- Rational optimization and fragment deconstruction can yield potent small molecule modulators.
- This study provides a foundation for developing SLIT2/ROBO1 pathway inhibitors for therapeutic applications.
