Small Molecule Inhibitors of Ca²⁺-S100B Reveal Two Protein Conformations

The drug pentamidine inhibits calcium-dependent complex formation with p53 (^CaS100B·p53) in malignant melanoma (MM) and restores p53 tumor suppressor activity in vivo. However, off-target effects associated with this drug were problematic in MM patients. Structure–activity relationship (SAR) studies were therefore completed here with 23 pentamidine analogues, and X-ray structures of ^CaS100B·inhibitor complexes revealed that the C-terminus of S100B adopts two different conformations, with location of Phe87 and Phe88 being the distinguishing feature and termed the “FF-gate”. For symmetric pentamidine analogues (^CaS100B·5a, ^CaS100B·6b) a channel between sites 1 and 2 on S100B was occluded by residue Phe88, but for an asymmetric pentamidine analogue (^CaS100B·17), this same channel was open. The ^CaS100B·17 structure illustrates, for the first time, a pentamidine analog capable of binding the “open” form of the “FF-gate” and provides a means to block all three “hot spots” on ^CaS100B, which will impact next generation ^CaS100B·p53 inhibitor design.