EphA4 targeting agents protect motor neurons from cell death induced by amyotrophic lateral sclerosis -astrocytes

Mixture-Based Screening of Focused Combinatorial Libraries by NMR: Application to the Antiapoptotic Protein hMcl-1

“…While the identification of initial weakly binding agents to a protein target requires sensitive biophysical approaches, such as NMR spectroscopy, these methods tend to usually present inherently a low throughput compared to typical HTS biochemical screens. To ameliorate this issue and increase the chemical space that can be sampled by NMR, we have recently proposed to prepare compound libraries in mixtures, arranged in a positional scanning (POS) fashion. − , Alternatively, we also proposed to derivatize a POS library with a common binding element for a given target or a class of targets. ,,, Hits within these peptide-like libraries still require significant optimization to obtain potent and selective druglike molecules. , To exemplify the library synthesis and the optimization of the resulting hits into more druglike molecules, we propose a new library and an associated deconvolution strategy. To reduce the peptide-like nature of the agents, the first element of the library was chosen among several sulfonyl chlorides that was efficiently reacted with the N-terminus of each element of a mixture of dipeptides (36 × 36 chosen among natural and non-natural aminoamides) using conventional solid-phase synthesis.…”

“…As mentioned above, to target PPIs, we have recently reported the HTS by NMR approach , that consists of combining principles of positional scanning combinatorial chemistry and protein-based NMR screening to identify, within libraries of >100,000 compounds, possible initial binders to protein–protein interactions. More recently, we also proposed deploying a “focused” positional scanned library ( f HTS by NMR) for the approach, in which each element of the library is designed to be derivatized with a target-specific (or a target class-specific) chemical binding moiety. ,, Using the HTS by NMR against EphA4-LBD, we identified initial ligands of weak affinity (>300 μM) that, nonetheless, using either the f HTS-by-NMR and/or iterative SAR optimizations, including the X-ray structure of the complex, could be optimized into potent and selective, low nanomolar affinity agents. ,,− Hence, deriving initial bona fide binding agents (as detected by sensitive protein-based NMR ligand binding assays) represents an exceptionally valuable starting point for the design of potent agents, especially if later supported by high-resolution structural information of the complex.…”

“…After SAR studies on compound 12 (Table S3), we thought to further enhance the binding affinity of the compound by elongating the molecule at the C-terminus with a fourth element. We proved this strategy recently in the design of potent agents targeting other PPIs involving the receptor tyrosine kinase EphA4 and its ephrin ligand. ,, …”

“… 15 , 16 , 22 , 23 Hits within these peptide-like libraries still require significant optimization to obtain potent and selective druglike molecules. 22 , 24 To exemplify the library synthesis and the optimization of the resulting hits into more druglike molecules, we propose a new library and an associated deconvolution strategy. To reduce the peptide-like nature of the agents, the first element of the library was chosen among several sulfonyl chlorides that was efficiently reacted with the N-terminus of each element of a mixture of dipeptides (36 × 36 chosen among natural and non-natural aminoamides) using conventional solid-phase synthesis.…”

“… 22 , 23 , 36 Using the HTS by NMR against EphA4-LBD, we identified initial ligands of weak affinity (>300 μM) that, nonetheless, using either the f HTS-by-NMR and/or iterative SAR optimizations, including the X-ray structure of the complex, could be optimized into potent and selective, low nanomolar affinity agents. 15 , 16 , 22 − 24 Hence, deriving initial bona fide binding agents (as detected by sensitive protein-based NMR ligand binding assays) represents an exceptionally valuable starting point for the design of potent agents, especially if later supported by high-resolution structural information of the complex.…”