Macrophage infectivity potentiator (Mip) and Mip-like
proteins are
virulence factors in a wide range of pathogens including Legionella pneumophila. These
proteins belong to the
FK506 binding protein (
FKBP) family of peptidyl-prolyl-
cis/trans-isomerases (PPIases). In L. pneumophila, the
PPIase activity of Mip is required for invasion of macrophages, transmigration through an in vitro lung-epithelial barrier, and full virulence in the guinea pig
infection model. Additionally, Mip is a moonlighting
protein that binds to
collagen IV in the extracellular matrix. Here, we describe the development and synthesis of
cycloheximide derivatives with adamantyl moieties as novel
FKBP ligands, and analyze their effect on the viability of L. pneumophila and other bacteria. All compounds efficiently inhibited
PPIase activity of the prototypic human
FKBP12 as well as Mip with IC50-values as low as 180 nM and 1.7 μM, respectively. Five of these derivatives inhibited the growth of L. pneumophila at concentrations of 30-40 μM, but exhibited no effect on other tested bacterial species indicating a specific spectrum of antibacterial activity. The derivatives carrying a 3,5-dimethyladamantan-1-[yl]
acetamide substitution (MT_30.32), and a 3-ethyladamantan-1-[yl]
acetamide substitution (MT_30.51) had the strongest effects in
PPIase- and liquid growth assays. MT_30.32 and MT_30.51 were also inhibitory in macrophage
infection studies without being cytotoxic. Accordingly, by applying a combinatorial approach, we were able to generate novel, hybrid inhibitors consisting of
cycloheximide and
adamantane, two known
FKBP inhibitors that interact with different parts of the
PPIase domain, respectively. Interestingly, despite the proven Mip-inhibitory activity, the viability of a Mip-deficient strain was affected to the same degree as its wild type. Hence, we also propose that
cycloheximide derivatives with adamantyl moieties are potent
PPIase inhibitors with multiple targets in L. pneumophila.