Lysine-selective molecular tweezers are cell penetrant and concentrate in lysosomes

Lysine-selective molecular tweezers are promising drug candidates against proteinopathies, viral infection, and bacterial biofilm. Despite demonstration of their efficacy in multiple cellular and animal models, important questions regarding their mechanism of action, including cell penetrance and intracellular distribution, have not been answered to date. The main impediment to answering these questions has been the low intrinsic fluorescence of the main compound tested to date, called CLR01. Here, we address these questions using new fluorescently labeled molecular tweezers derivatives. We show that these compounds are internalized in neurons and astrocytes, at least partially through dynamin-dependent endocytosis. In addition, we demonstrate that the molecular tweezers concentrate rapidly in acidic compartments, primarily lysosomes. Accumulation of molecular tweezers in lysosomes may occur both through the endosomal-lysosomal pathway and via the autophagy-lysosome pathway. Moreover, by visualizing colocalization of molecular tweezers, lysosomes, and tau aggregates we show that lysosomes likely are the main site for the intracellular anti-amyloid activity of molecular tweezers. These findings have important implications for the mechanism of action of molecular tweezers in vivo, explaining how administration of low doses of the compounds achieves high effective concentrations where they are needed, and supporting the development of these compounds as drugs for currently cureless proteinopathies.

Optical Sensing and Supramolecular Cyanide Recognition Sites of Salophenes as Molecular Tweezers: The First Example of Recognition via Intramolecular Aldimine Condensation Cyclization

The optical sensing and supramolecular cyanide recognition sites of two dipodal Schiff bases having conjugated to catecols (1) and phenols (2) (namely, salophenes 1 and 2) have been studied. In the case of optical sensing, both probes recognized only cyanide (CN¯) ions in 30% PBS buffer CH3CN (pH 7.4) as confirmed from the changes of absorption and emission bands, resulting in color changes. From the supramolecular recognition point of view, probes 1 and 2 show the different recognition bahaviour toward CN¯, as evidenced by the fluoroscence and NMR data, as well the OH¯ and reversibility experiments. While 1 recognizes CN¯ via deprotonation, that of 2 is the first example of Schiff base which senses CN¯ through an intramolecular aldimine condensation cyclization, leading to formation of dihydroxyquinoxaline 4. In general, probes 1, 2 and 4 are promising on-site optical sensors in terms of easy prepared, selectivity, sensitivity (1–10 nM), ease of use, rapid response (< 5 s) and test kits.

Specific inhibition of the Survivin–CRM1 interaction by peptide-modified molecular tweezers

Survivin’s dual function as apoptosis inhibitor and regulator of cell proliferation is mediated via its interaction with the export receptor CRM1. This protein–protein interaction represents an attractive target in cancer research and therapy. Here, we report a sophisticated strategy addressing Survivin’s nuclear export signal (NES), the binding site of CRM1, with advanced supramolecular tweezers for lysine and arginine. These were covalently connected to small peptides resembling the natural, self-complementary dimer interface which largely overlaps with the NES. Several biochemical methods demonstrated sequence-selective NES recognition and interference with the critical receptor interaction. These data were strongly supported by molecular dynamics simulations and multiscale computational studies. Rational design of lysine tweezers equipped with a peptidic recognition element thus allowed to address a previously unapproachable protein surface area. As an experimental proof-of-principle for specific transport signal interference, this concept should be transferable to any protein epitope with a flanking well-accessible lysine.

Lysine-selective molecular tweezers are cell-penetrant and concentrate in lysosomes

Lysine-selective molecular tweezers are promising drug candidates against proteinopathies and viral infection. Despite demonstration of their efficacy in multiple cellular and animal models, important questions regarding their mechanism of action, including cell penetrance and intracellular distribution, have not been answered to date. The main impediment to answering these questions has been the low intrinsic fluorescence of the main compound tested to date, called CLR01. Here, we address these questions using new fluorescently labeled molecular tweezers derivatives. We show that these compounds are internalized in neurons and astrocytes, at least partially through dynamin-dependent endocytosis. In addition, we demonstrate that the molecular tweezers concentrate rapidly in acidic compartments, primarily lysosomes. Accumulation of molecular tweezers in lysosomes may occur both through the endosomal-lysosomal pathway and via the autophagy-lysosome pathway. Moreover, by visualizing colocalization of molecular tweezers, lysosomes, and tau aggregates we show that lysosomes likely are the main site for the intracellular anti-amyloid activity of molecular tweezers. These findings have important implications for the mechanism of action of molecular tweezers in vivo, explaining how administration of low doses of the compounds achieves high effective concentrations where they are needed, and supporting the development of these compounds as drugs for currently cureless proteinopathies.

ON/OFF metal-triggered molecular tweezers for fullerene recognition

Herein, we report molecular tweezers for fullerene recognition based on 2,2’-bipyridine-bearing corannulene motifs. The syn or anti confirmation can be selected simply by Cu(I) coordination/decoordination, thus controlling the fullerene recognition...