Ulrike Holzgrabe

Due to increasing antibiotic resistance, the development of anti‐infectives with new mechanisms of action is crucial. Virulence factors such as the “macrophage infectivity potentiator” (Mip) protein, which catalyzes the folding of proline‐containing proteins by means of their cis–trans isomerase (PPIase) activity, have come into focus as a potential new target. Since the inhibition of Mip by small molecules has been shown to lead to reduced virulence and survival in vitro, especially of Gram‐negative bacteria such as Burkholderia pseudomallei (Bp), Neisseria meningitidis (Nm), and Neisseria gonorrhoeae (Ng), or Coxiella burnetii (Cb), among many others, a library of Mip inhibitors was developed. As drug metabolism has a significant impact on the overall therapeutic outcome, this report describes the biotransformation of the most potent Mip inhibitors. Therefore, the anti‐infectives were treated using human liver microsomes in vitro. Liquid chromatography with tandem mass spectrometry (LC/MS‐MS) methods were applied to identify the metabolites and quantify the metabolic degradation of the hit compounds. Active metabolites, N ‐oxides, were found, leading to new opportunities for further drug development.

In the last few years, fluorescence resonance energy transfer (FRET) receptor sensors have contributed to the understanding of GPCR ligand binding and functional activation. FRET sensors based on muscarinic acetylcholine receptors (mAChRs) have been employed to study dual-steric ligands, allowing for the detection of different kinetics and distinguishing between partial, full, and super agonism. Herein, we report the synthesis of the two series of bitopic ligands, 12-Cn and 13-Cn, and their pharmacological investigation at the M1, M2, M4, and M5 FRET-based receptor sensors. The hybrids were prepared by merging the pharmacophoric moieties of the M1/M4-preferring orthosteric agonist Xanomeline 10 and the M1-selective positive allosteric modulator 77-LH-28-1 (1-[3-(4-butyl-1-piperidinyl)propyl]-3,4-dihydro-2(1H)-quinolinone) 11. The two pharmacophores were connected through alkylene chains of different lengths (C3, C5, C7, and C9). Analyzing the FRET responses, the tertiary amine compounds 12-C5, 12-C7, and 12-C9 evidenced a selective activation of M1 mAChRs, while the methyl tetrahydropyridinium salts 13-C5, 13-C7, and 13-C9 showed a degree of selectivity for M1 and M4 mAChRs. Moreover, whereas hybrids 12-Cn showed an almost linear response at the M1 subtype, hybrids 13-Cn evidenced a bell-shaped activation response. This different activation pattern suggests that the positive charge anchoring the compound 13-Cn to the orthosteric site ensues a degree of receptor activation depending on the linker length, which induces a graded conformational interference with the binding pocket closure. These bitopic derivatives represent novel pharmacological tools for a better understanding of ligand-receptor interactions at a molecular level.