Discovery, Structure-Activity Relationship Studies, and Crystal Structure of Nonpeptide Inhibitors Bound to the Shank3 PDZ Domain

• Published in: ChemMedChem 2011-08, Vol.6 (8), p.1411-1422
• Main Authors: Saupe, Jörn, Roske, Yvette, Schillinger, Christian, Kamdem, Nestor, Radetzki, Silke, Diehl, Anne, Oschkinat, Hartmut, Krause, Gerd, Heinemann, Udo, Rademann, Jörg
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Binding Sites; Carrier Proteins - chemistry; Carrier Proteins - genetics; Carrier Proteins - metabolism; Crystallography, X-Ray; Drug Evaluation, Preclinical; fluorescence polarization; high-throughput screening; Humans; inhibitors; Nerve Tissue Proteins; PDZ domain; PDZ Domains - drug effects; Peptides - chemical synthesis; Peptides - chemistry; Protein Interaction Domains and Motifs; protein-protein interactions; Quinolines - chemical synthesis; Quinolines - chemistry; Quinolines - pharmacology; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Structure-Activity Relationship
• ISSN: 1860-7179; 1860-7187
• DOI: 10.1002/cmdc.201100094

Shank is the central scaffolding protein of the postsynaptic density (PSD) protein complex found in cells of the central nervous system. Cellular studies indicate a prominent role of the protein in the organization of the PSD, in the development of neuronal morphology, in neuronal signaling, and in synaptic plasticity, thus linking Shank functions to the molecular basis of learning and memory. Mutations in the Shank gene have been found in several neuronal disorders including mental retardation, typical autism, and Asperger syndrome. Shank is linked to the PSD complex via its PDZ domain that binds to the C‐terminus of guanylate‐kinase‐associated protein (GKAP). Here, small‐molecule inhibitors of Shank3 PDZ domain are developed. A fluorescence polarization assay based on an identified high‐affinity peptide is established, and tetrahydroquinoline carboxylates are identified as inhibitors of this protein–protein interaction. Chemical synthesis via a hetero‐Diels–Alder strategy is employed for hit optimization, and structure–activity relationship studies are performed. Best hits possess Ki values in the 10 μM range, and binding to the PDZ domain is confirmed by 1H,15N HSQC NMR experiments. One of the hits crystallizes with the Shank3 PDZ domain. The structure, analyzed at a resolution of 1.85 Å, reveals details of the binding mode. Finally, binding to PDZ domains of PSD‐95, syntrophin, and DVL3 was studied using 1H,15N HSQC NMR spectroscopy. Shank in the crosshairs: We discovered inhibitors of the Shank3 PDZ domain, a putative target in autism spectrum disorders. Tetrahydroquinoline carboxylates were found by HTS using a fluorescence polarization assay and represent a new class of PDZ domain ligands. The crystal structure of the Shank3 PDZ domain in complex with a tetrahydroquinoline derivative reveals details of the binding mode.