Synthesis of Bis(1,2,3-Triazole) Functionalized Quinoline-2,4-Diones.

Derivatives of 3-(1H-1,2,3-triazol-1-yl)quinoline-2,4(1H,3H)-dione unsubstituted on quinolone nitrogen atom, which are available by the previously described four step synthesis starting from aniline, were exploited as intermediates in obtaining the title compounds. The procedure involves the introduction of propargyl group onto the quinolone nitrogen atom of mentioned intermediates by the reaction of them with propargyl bromide in N,N-dimethylformamide (DMF) in presence of a potassium carbonate and the subsequent formation of a second triazole ring by copper catalyzed cyclisation reaction with azido compounds. The products were characterized by ¹H, 13C and 15N NMR spectroscopy. The corresponding resonances were assigned on the basis of the standard 1D and gradient selected 2D NMR experiments (¹H⁻¹H gs-COSY, ¹H⁻13C gs-HSQC, ¹H⁻13C gs-HMBC) with ¹H⁻15N gs-HMBC as a practical tool to determine 15N NMR chemical shifts at the natural abundance level of 15N isotope.

Novel Peptide-Based Fluorescent Probe for Simultaneous Sensing of Chymotrypsin and Hydrogen Peroxide.

The developed multifunctional fluorescent probe enables the simultaneous detection of chymotrypsin as a model protease and hydrogen peroxide as a representative of reactive oxygen species (ROS) in biologically relevant concentration ranges. The chymotrypsin sensing is based on the cleavage of its selectively recognizable peptide sequence and the consequent disruption of FRET between coumarin (DEAC) and fluorescein (FL). Analogously, the presence of hydrogen peroxide causes the gradual degradation of the H2O2-labile benzopyrylium-coumarin (BC) dye. Considering the fluorescence emission responses of individual chymotrypsin-peroxide probe-attached fluorophores after their excitation at 425 nm, the sole presence of either chymotrypsin (50-1000 ng/mL) or hydrogen peroxide (10-200 µM) in a sample could be unambiguously confirmed or refuted. In addition, reliable simultaneous detection and approximate quantification of both studied species in the concentration ranges of 100-1000 ng/mL and 20-200 µM for chymotrypsin and H2O2, respectively, could be performed as well. The obtained results are summarized and visualized in the graphical models.

Triple-FRET multi-purpose fluorescent probe for three-protease detection.

A new, robust and reliable methodology for three-protease screening in a single-enzyme mode has been developed and verified, employing a multi-purpose peptide probe with three selectively cleavable sites furnished with four fluorophores. A triple-FRET-based single-excitation quadruple-emission concept for unambiguous sensing of trypsin, chymotrypsin and caspase-8 in the lowest detectable concentrations of 0.5 ng mL-1, 0.2 µg mL-1, and 2 U mL-1, respectively, has been applied and graphically depicted. Then the developed 4-dye probe has been also studied from the perspective of simultaneous two-protease screening, which was found only partially feasible, primarily due to unselective chymotrypsin cleavage.

Cytotoxicity of Amino-BODIPY Modulated via Conjugation with 2-Phenyl-3-Hydroxy-4(1H)-Quinolinones.

The combination of cytotoxic amino-BODIPY dye and 2-phenyl-3-hydroxy-4(1H)-quinolinone (3-HQ) derivatives into one molecule gave rise to selective activity against lymphoblastic or myeloid leukemia and the simultaneous disappearance of the cytotoxicity against normal cells. Both species' conjugation can be realized via a disulfide linker cleavable in the presence of glutathione characteristic for cancer cells. The cleavage liberating the free amino-BODIPY dye and 3-HQ derivative can be monitored by ratiometric fluorescence or by the OFF-ON effect of the amino-BODIPY dye. A similar cytotoxic activity is observed when the amino-BODIPY dye and 3-HQ derivative are connected through a non-cleavable maleimide linker. The work reports the synthesis of several conjugates, the study of their cleavage inside cells, and cytotoxic screening.

Preparation of Quinoline-2,4-dione Functionalized 1,2,3-Triazol-4-ylmethanols, 1,2,3-Triazole-4-carbaldehydes and 1,2,3-Triazole-4-carboxylic Acids.

(1-(2,4-Dioxo-1,2,3,4-tetrahydroquinolin-3-yl)-1H-1,2,3-triazol-4-yl)methyl acetates substituted on nitrogen atom of quinolinedione moiety with propargyl group or (1-substituted 1H-1,2,3-triazol-4-yl)methyl group, which are available from the appropriate 3-(4-hydroxymethyl-1H-1,2,3-triazol-1-yl)quinoline-2,4(1H,3H)-diones unsubstituted on quinolone nitrogen atom by the previously described procedures, were deacetylated by acidic ethanolysis. Thus obtained primary alcohols, as well as those aforenamed unsubstituted on quinolone nitrogen atom, were oxidized to aldehydes on the one hand with pyridinium chlorochromate (PCC), on the other hand with manganese dioxide, and to carboxylic acids using Jones reagent in acetone. The structures of all prepared compounds were confirmed by 1H, 13C and 15N NMR spectroscopy. The corresponding resonances were assigned on the basis of the standard 1D and gradient selected 2D NMR experiments (1H-1H gs-COSY, 1H-13C gs-HSQC, 1H-13C gs-HMBC) with 1H-15N gs-HMBC as a practical tool to determine 15N NMR chemical shifts at the natural abundance level of 15N isotope.

Design, synthesis and antitubercular potency of 4-hydroxyquinolin-2(1H)-ones.

In this study, a 50-membered library of substituted 4-hydroxyquinolin-2(1H)-ones and two closely related analogues was designed, scored in-silico for drug likeness and subsequently synthesized. Thirteen derivatives, all sharing a common 3-phenyl substituent showed minimal inhibitory concentrations against Mycobacterium tuberculosis H37Ra below 10 µM and against Mycobacterium bovis AN5A below 15 µM but were inactive against faster growing mycobacterial species. None of these selected derivatives showed significant acute toxicity against MRC-5 cells or early signs of genotoxicity in the Vitotox™ assay at the active concentration range. The structure activity study relation provided some insight in the further favourable substitution pattern at the 4-hydroxyquinolin-2(1H)-one scaffold and finally 6-fluoro-4-hydroxy-3-phenylquinolin-2(1H)-one (38) was selected as the most promising member of the library with a MIC of 3.2 µM and a CC50 against MRC-5 of 67.4 µM.