A three-stage biophysical screening cascade for fragment-based drug discovery

Area of Science:

• Biochemistry
• Structural Biology
• Drug Discovery

Background:

• Fragment-based drug discovery (FBDD) is a key strategy for developing high-affinity ligands.
• Sensitive biophysical techniques are crucial for detecting weak interactions between small molecules and target macromolecules.
• Identifying initial weak binders is essential for designing potent inhibitors.

Purpose of the Study:

• To describe an efficient protocol for screening compound libraries using biophysical ligand-binding assays.
• To outline a multi-stage cascade for identifying and validating binding fragments (hits).
• To provide a robust method applicable to both academic and industrial drug discovery settings.

Main Methods:

• Utilizing a cascade approach involving differential scanning fluorimetry (DSF) for preliminary screening.
• Employing NMR spectroscopy for validation of potential hits.
• Characterizing binding fragments using isothermal titration calorimetry (ITC) and X-ray crystallography.

Main Results:

• An efficient fragment screening cascade capable of identifying binding fragments within weeks.
• Demonstration of orthogonal validation using complementary biophysical techniques.
• Establishment of a protocol suitable for low-cost, medium-throughput early-stage screening.

Conclusions:

• The described fragment screening cascade offers an efficient route to identify starting points for drug development.
• The protocol's emphasis on orthogonal validation ensures robustness and quality suitable for industrial applications.
• This method facilitates the design of high-affinity ligands for therapeutic targets.