title: Covalent small molecule inhibitors of Ca(2+)-bound S100B.
abstract: Elevated levels of the tumor marker S100B are observed in malignant melanoma, and this EF-hand-containing protein was shown to directly bind wild-type (wt) p53 in a Ca(2+)-dependent manner, dissociate the p53 tetramer, and inhibit its tumor suppression functions. Likewise, inhibiting S100B with small interfering RNA (siRNA(S100B)) is sufficient to restore wild-type p53 levels and its downstream gene products and induce the arrest of cell growth and UV-dependent apoptosis in malignant melanoma. Therefore, it is a goal to develop S100B inhibitors (SBiXs) that inhibit the S100B-p53 complex and restore active p53 in this deadly cancer. Using a structure-activity relationship by nuclear magnetic resonance approach (SAR by NMR), three persistent binding pockets are found on S100B, termed sites 1-3. While inhibitors that simultaneously bind sites 2 and 3 are in place, no molecules that simultaneously bind all three persistent sites are available. For this purpose, Cys84 was used in this study as a potential means to bridge sites 1 and 2 because it is located in a small crevice between these two deeper pockets on the protein. Using a fluorescence polarization competition assay, several Cys84-modified S100B complexes were identified and examined further. For five such SBiX-S100B complexes, crystallographic structures confirmed their covalent binding to Cys84 near site 2 and thus present straightforward chemical biology strategies for bridging sites 1 and 3. Importantly, one such compound, SC1982, showed an S100B-dependent death response in assays with WM115 malignant melanoma cells, so it will be particularly useful for the design of SBiX molecules with improved affinity and specificity.
authors.affiliation: Center for Biomolecular Therapeutics (CBT), Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine , Baltimore, Maryland 21201, United States.
authors.forename: Michael C
authors.initials: MC
authors.lastname: Cavalier
authors.forename: Adam D
authors.initials: AD
authors.lastname: Pierce
authors.forename: Paul T
authors.initials: PT
authors.lastname: Wilder
authors.forename: Milad J
authors.initials: MJ
authors.lastname: Alasady
authors.forename: Kira G
authors.initials: KG
authors.lastname: Hartman
authors.forename: David B
authors.initials: DB
authors.lastname: Neau
authors.forename: Timothy L
authors.initials: TL
authors.lastname: Foley
authors.forename: Ajit
authors.initials: A
authors.lastname: Jadhav
authors.forename: David J
authors.initials: DJ
authors.lastname: Maloney
authors.forename: Anton
authors.initials: A
authors.lastname: Simeonov
authors.forename: Eric A
authors.initials: EA
authors.lastname: Toth
authors.forename: David J
authors.initials: DJ
authors.lastname: Weber
mesh_terms: D000818:Animals; D053119:Benzophenanthridines / Q000737:chemistry / Q000494:pharmacology; D016227:Benzoquinones / Q000737:chemistry / Q000494:pharmacology; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002413:Cations, Divalent; D002417:Cattle; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018360:Crystallography, X-Ray; D004221:Disulfiram / Q000737:chemistry / Q000494:pharmacology; D004224:Diterpenes / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D008545:Melanoma; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D064568:S100 Calcium Binding Protein beta Subunit / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism
chemical_list: D053119:Benzophenanthridines; D016227:Benzoquinones; D002413:Cations, Divalent; D004224:Diterpenes; D064568:S100 Calcium Binding Protein beta Subunit; C026197:3,4-dimethoxydalbergione; C038061:rubescensine A; C016299:chelerythrine; D002118:Calcium; D004221:Disulfiram