Probing pockets S2–S4′ of the γ-secretase active site with (hydroxyethyl)urea peptidomimetics

(Hydroxyethyl)urea peptidomimetics are potent inhibitors of γ-secretase that are accessible in a few synthetic steps. Systematic alteration of P2–P4′ revealed that the corresponding S2–S4′ active site pockets accommodate a variety of substituents, consistent with the fact that this protease cleaves a variety of single-pass membrane proteins; however, phenylalanine is not well tolerated at P2′. A compound spanning P2–P3′ was identified as a low nM inhibitor of γ-secretase activity both in cells and under cell-free conditions.