Multi-targeting protein-protein interaction inhibitors: Evolution of macrocyclic ligands with embedded carbohydrates (MECs) to improve selectivity.

Compounds targeting multiple proteins can have synergistic effects and are therefore of interest in medicinal chemistry. At the same time, inhibiting protein-protein interactions (PPI) is increasingly desired in the treatment of disorders or diseases. The development of non-peptidomimetic inhibitors is still a challenge. Herein we investigate macrocyclic scaffolds with one or two embedded carbohydrates (MECs) that present amino acid side chains, or related isosteres, as pharmacophoric groups. Firstly, retroscreening of the previously reported eannaphane-40 (E40, 40), a MEC presenting two pharmacophoric groups, against a set of 55 receptor-subtypes led to a finding of sub-micromolar inhibitory activity for E40 against three serotonergic isoforms (5HT1A/2A/2B) as well as the Na+ channel and the NK-2 receptor. We synthesised MECs with an additional pharmacophoric group compared to E40, with a view to identifying compounds where the selectivity profile was altered among the protein hits from the retroscreening. MECs were produced based on scaffolds with two monosaccharide residues, leading to the incorporation of a third pharmacophoric group. Later, homology models were prepared for four proteins (5HT1A, 5HT2A, NK2 and site-2 of the sodium channel) whose 3D structure is unknown. Inverse docking of the synthesised compounds led to the selection of a new MEC (MEC-B) for protein binding assays. MEC-B was found to have its selectivity profile modulated, in line with docking prediction, compared to E40. MEC-B is dual inhibitor of both 5-HT1A and the sodium channel with improved selectivity for these proteins compared to 5-HT2A/2B/2C, 5-HT transporter and NK2 receptor. Thus, a new multitargeting compound, with an improved selectivity profile was identified, based on a MEC peptidomimetic scaffold.

Decorated macrocycles via ring-closing double-reductive amination. identification of an apoptosis inducer of leukemic cells that at least partially antagonizes a 5-HT2 receptor.

A build-couple-pair strategy, including double-reductive amination macrocyclization, has been used to generate decorated macrocycles (eannaphanes) with an embedded triazole and monosaccharide. Biological screening led to the identification of an inducer of apoptosis in leukemic cells, which acts at least partially as a 5-HT2 antagonist.

Synthesis of bivalent lactosides and their activity as sensors for differences between lectins in inter- and intrafamily comparisons.

The synthesis of nine bivalent lactosides (based on ditriazoles, diamides, a glycocyclophane and an acyclic analogue of the glycocyclophane) and one monovalent lactosyl triazole facilitated the assessment of the sensitivity of plant/animal lectins to this type of ligand display. The inhibitory potency of the compounds was determined in two assays of increasing biorelevance. These were solid-phase and cell binding set-ups. Hereby, the ability of the compounds to inhibit the binding of two plant agglutinins and the entire set of adhesion/growth-regulatory galectins from one organism (chicken) to a glycoprotein or to cell surfaces was systematically evaluated. Differential sensitivities were detected between plant and animal lectins and also between distinct galectin forms within the chicken series. Two of the bivalent probes can be considered as sensors for interlectin differences. Most pronounced were the selectivities of N-glycosyl 1,2,3-triazole derivatives for the chimera-type galectin and its proteolytically truncated version.

Towards echinomycin mimetics by grafting quinoxaline residues on glycophane scaffolds.

Echinomycin is a natural depsipeptide, which is a bisintercalator, inserting quinoxaline units preferentially adjacent to CG base pairs of DNA. Herein the design and synthesis of echinomycin mimetics based on grafting of two quinoxaline residues onto a macrocyclic scaffold (glycophane) is addressed. Binding of the compounds to calf-thymus DNA was studied using UV-vis and steady state fluorescence spectroscopy, as well as thermal denaturation. An interesting observation was enhancement of fluorescence emission for the peptidomimetics on binding to DNA, which contrasted with observations for echinomycin. Molecular dynamics simulations were exploited to explore in more detail if bis-intercalation to DNA was possible for one of the glycophanes. Bis-intercalating echinomycin complexes with DNA were found to be stable during 20ns simulations at 298K. However, the MD simulations of a glycophane complexed with a DNA octamer displayed very different behaviour to echinomycin and its quinoxaline units were found to rapidly migrate out from the intercalation site. Release of bis-intercalation strain occurred with only one of the quinoxaline chromophores remaining intercalated throughout the simulation. The distance between the quinoxaline residues in the glycophane at the end of the MD simulation was 7.3-7.5Å, whereas in echinomycin, the distance between the residues was ∼11Å, suggesting that longer glycophane scaffolds would be required to generate bis-intercalating echinomycin mimetics.

FIT probes: peptide nucleic acid probes with a fluorescent base surrogate enable real-time DNA quantification and single nucleotide polymorphism discovery.

The ability to accurately quantify specific nucleic acid molecules in complex biomolecule solutions in real time is important in diagnostic and basic research. Here we describe a DNA-PNA (peptide nucleic acid) hybridization assay that allows sensitive quantification of specific nucleic acids in solution and concomitant detection of select single base mutations in resulting DNA-PNA duplexes. The technique employs so-called FIT (forced intercalation) probes in which one base is replaced by a thiazole orange (TO) dye molecule. If a DNA molecule that is complementary to the FIT-PNA molecule (except at the site of the dye) hybridizes to the probe, the TO dye exhibits intense fluorescence because stacking in the duplexes enforces a coplanar arrangement even in the excited state. However, a base mismatch at either position immediately adjacent to the TO dye dramatically decreases fluorescence, presumably because the TO dye has room to undergo torsional motions that lead to rapid depletion of the excited state. Of note, we found that the use of d-ornithine rather than aminoethylglycine as the PNA backbone increases the intensity of fluorescence emitted by matched probe-target duplexes while specificity of fluorescence signaling under nonstringent conditions is also increased. The usefulness of the ornithine-containing FIT probes was demonstrated in the real-time PCR analysis providing a linear measurement range over at least seven orders of magnitude. The analysis of two important single nucleotide polymorphisms (SNPs) in the CFTR gene confirmed the ability of FIT probes to facilitate unambiguous SNP calls for genomic DNA by quantitative PCR.

Ultrasound promoted Suzuki cross-coupling reactions in ionic liquid at ambient conditions.

Palladium catalyzed Suzuki cross-coupling reactions of halobenzenes including chlorobenzenes with phenylboronic acid have been achieved at ambient temperature (30 degrees C) in the absence of a phosphine ligand using the ionic liquid 1,3-di-n-butylimidazolium tetrafluoroborate [bbim][BF4] with methanol as co-solvent under ultrasonic irradiation.