Mono- and bis(steroids) containing a cyclooctane core: Synthesis, antiproliferative activity, and action on cell cytoskeleton microtubules.

Steroid dimers of natural and synthetic origin possess an unusual and complex molecular architecture that may lead to the realization of peculiar effects in biological systems, in particular in different cancer cell lines. In the present work, diastereoselective ring-opening of mono- and polyoxiranes, containing a cyclooctane core, by azide-anion was performed to yield a series of azidoalcohols with different types of symmetry. The products were involved in copper-catalyzed azyde-alkyne cycloaddition (CuAAC) reaction with ethinylestradiol and ethinyltestosterone, and the resulting steroids and steroid dimers with triazole linkers were screened for their antiproliferative activity via (3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide) assay. All the compounds revealed cytotoxicity toward several cancer cell lines. The effect of the most potent compound, containing two estradiol moieties, on the microtubules (MT) dynamics was investigated by immunofluorescent microscopy. The disruption of the majority of interphase cell cytoplasmic MT and mitotic event disturbances in the presence of the studied compound were observed. The latter effect caused the appearance of numerous multinucleated cells.

Verubulin (Azixa) Analogues with Increased Saturation: Synthesis, SAR and Encapsulation in Biocompatible Nanocontainers Based on Ca2+ or Mg2+ Cross-Linked Alginate.

Tubulin-targeting agents attract undiminished attention as promising compounds for the design of anti-cancer drugs. Verubulin is a potent tubulin polymerization inhibitor, binding to colchicine-binding sites. In the present work, a series of verubulin analogues containing a cyclohexane or cycloheptane ring 1,2-annulated with pyrimidine moiety and various substituents in positions 2 and 4 of pyrimidine were obtained and their cytotoxicity towards cancer and non-cancerous cell lines was estimated. The investigated compounds revealed activity against various cancer cell lines with IC50 down to 1-4 nM. According to fluorescent microscopy data, compounds that showed cytotoxicity in the MTT test disrupt the normal cytoskeleton of the cell in a pattern similar to that for combretastatin A-4. The hit compound (N-(4-methoxyphenyl)-N,2-dimethyl-5,6,7,8-tetrahydroquinazolin-4-amine) was encapsulated in biocompatible nanocontainers based on Ca2+ or Mg2+ cross-linked alginate and it was demonstrated that its cytotoxic activity was preserved after encapsulation.

A new oxidatively stable ligand for the chiral functionalization of amino acids in Ni(II)-Schiff base complexes.

A new oxidatively stable (S)-N-benzylproline-derived ligand ((S)-N-(2-benzoyl-5-tert-butylphenyl)-1-benzylpyrrolidine-2-carboxamide) and its Ni(II)-Schiff base complexes formed of glycine, serine, and dehydroalanine are reported. A bulky tert-butyl substituent in the phenylene fragment precludes unwanted oxidative dimerization of the Schiff base complex, making it suitable for targeted electrochemically induced oxidative modification of the amino acid side chain. Experimental and DFT studies showed that the additional tert-butyl group increases the dispersion interactions in the Ni coordination environment making the complexes more conformationally rigid and provides a higher level of thermodynamically controlled stereoselectivity as compared to the parent Belokon complex. Additionally, functionalization with the tert-butyl group significantly enhances the reactivity of the deprotonated glycine complex towards electrophiles as compared to the anionic species formed from the original Belokon complex. Solubility of the t-Bu-containing ligand and its Schiff base complexes is increased, facilitating scaling-up the reaction procedure and isolation of the functionalized amino acid.

[3+2]-Cycloaddition of Nitrile Imines to Parabanic Acid Derivatives-An Approach to Novel Spiroimidazolidinediones.

Approximately 1,3-Dipolar cycloaddition of imidazolidine derivatives containing exocyclic double bonds is a convenient method of creating spiro-conjugated molecules with promising anticancer activity. In this work, the derivatives of parabanic acid (2-thioxoimidazolidine-4,5-diones and 5-aryliminoimidazolidine-2,4-diones) were first investigated as dipolarophiles in the reactions with nitrile imines. The generation of nitrile imines was carried out either by the addition of tertiary amine to hydrazonoyl chlorides «drop by drop¼ or using the recently proposed diffusion mixing technique, which led to ~5-15% increases in target compound yields. It was found that the addition of nitrile imines to C=S or C=N exocyclic double bonds led to 1,2,4-thiazolines or triazolines and occurred regioselectively in accordance with the ratio of FMO coefficients of reactants. The yield of the resulting spiro-compound depended on the presence of alkyl substituents in the nitrile imine structure and was significantly decreased in reactions with imines with strong electron donor or electron-withdrawing groups. Some of the obtained compounds showed reasonable in vitro cytotoxicity. IC50 values were calculated for HCT116 (colon cancer) cells using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test.

Reductive opening of a cyclopropane ring in the Ni(II) coordination environment: a route to functionalized dehydroalanine and cysteine derivatives.

The involvement of an α,α-cyclopropanated amino acid in the chiral Ni(II) coordination environment in the form of a Schiff base is considered as a route to electrochemical broadening of the donor-acceptor cyclopropane concept in combination with chirality induction in the targeted products. A tendency to the reductive ring-opening and the follow-up reaction paths of thus formed radical anions influenced by substituents in the cyclopropane ring are discussed. Optimization of the reaction conditions opens a route to the non-proteinogenic amino acid derivatives containing an α-ß or ß-γ double C=C bond in the side chain; the regioselectivity can be tuned by the addition of Lewis acids. One-pot combination of the reductive ring opening and subsequent addition of thiols allows obtaining the cysteine derivatives in practical yields and with high stereoselectivity at the removed ß-stereocenter.

Novel substituted 5-methyl-4-acylaminoisoxazoles as antimitotic agents: Evaluation of selectivity to LNCaP cancer cells.

A series of novel antimitotic agents was designed using the replacement of heterocyclic cores in two tubulin-targeting lead molecules with the acylated 4-aminoisoxazole moiety. Target compounds were synthesized via heterocyclization of ß-aryl-substituted vinylketones by tert-butyl nitrite in the presence of water as a key step. 4-Methyl-N-[5-methyl-3-(3,4,5-trimethoxyphenyl)isoxazol-4-yl]benzamide (1aa) was found to stimulate partial depolymerization of microtubules of human lung carcinoma A549 cells at a high concentration of 100 µM and to totally inhibit cell growth (IC50 = 0.99 µM) and cell viability (IC50 = 0.271 µM) in the nanomolar to submicromolar concentration range. These data provide evidence of the multitarget profile of the cytotoxic action of compound 1aa. The SAR study demonstrated that the 3,4,5-trimethoxyphenyl residue is the key structural parameter determining the efficiency both towards tubulin and other molecular targets. The cytotoxicity of 3-methyl-N-[5-methyl-3-(3,4,5-trimethoxyphenyl)isoxazol-4-yl]benzamide (1ab) to the androgen-sensitive human prostate adenocarcinoma cancer cell line LNCaP (IC50 = 0.301 µM) was approximately one order of magnitude higher than that to the conditionally normal cells lines WI-26 VA4 (IC50 = 2.26 µM) and human umbilical vein endothelial cells (IC50 = 5.58 µM) and significantly higher than that to primary fibroblasts (IC50 > 75 µM).

Polypeptide-Based Molecular Platform and Its Docetaxel/Sulfo-Cy5-Containing Conjugate for Targeted Delivery to Prostate Specific Membrane Antigen.

A strategy for stereoselective synthesis of molecular platform for targeted delivery of bimodal therapeutic or theranostic agents to the prostate-specific membrane antigen (PSMA) receptor was developed. The proposed platform contains a urea-based, PSMA-targeting Glu-Urea-Lys (EuK) fragment as a vector moiety and tripeptide linker with terminal amide and azide groups for subsequent addition of two different therapeutic and diagnostic agents. The optimal method for this molecular platform synthesis includes (a) solid-phase assembly of the polypeptide linker, (b) coupling of this linker with the vector fragment, (c) attachment of 3-aminopropylazide, and (d) amide and carboxylic groups deprotection. A bimodal theranostic conjugate of the proposed platform with a cytostatic drug (docetaxel) and a fluorescent label (Sulfo-Cy5) was synthesized to demonstrate its possible sequential conjugation with different functional molecules.

Speciation of structural fragments in crude oil by means of isotope exchange in near-critical water and Fourier transform mass spectrometry.

The structures of individual molecules in crude oil remain largely unknown despite the considerable amount of research dedicated to this topic. The extreme complexity of crude oil (recently Marshall reported the observation of more than 400,000 unique compounds in one sample) makes it impossible to separate crude oil into individual compounds and determine their structure by NMR or X-ray spectroscopy. Recently, isotope exchange, performed both in solution and in the gas phase, combined with high-resolution mass spectrometry was used for speciation of certain structural fragments of individual molecules in crude oil and humic substances. 16O/18O exchange allows enumeration of =O groups and speciation of furans, whereas H/D exchange allows enumeration of -OH groups, -NH groups, aromatic hydrogens, alpha hydrogens, etc. Unfortunately, crude oil is insoluble in water (the most available and cleanest source of isotopes), so performance of the exchange in solution requires harsh conditions, such as concentrated acids or bases, which could considerably modify the sample. Here we describe the use of a cheap and simple analytical approach for performing both H/D and 16O/18O exchange in crude oil using only water as the source of the isotopes. Crude oil was incubated in near-critical water and the reaction was monitored by high-resolution Fourier transform mass spectrometry. Although isotope exchange results in complication of the spectrum, the resolving power of modern mass spectrometers is sufficient to determine the number of exchanges for each molecule simultaneously. We determined the number of 16O/18O exchanges in 276 species and the number of H/D exchanges in 150 species. Our results allow deeper investigation of crude oil and other nonpolar samples on the molecular level. Graphical abstract.

Individual (f,t A)- and (f,t C)-Fullerene-Based Nickel(II) Glycinates: Protected Chiral Amino Acids Directly Linked to a Chiral π-Electron System.

Stereoselective electrosynthesis of the first individual (f,t A)- and (f,t C)-1,4-fullerene derivatives with a non-inherently chiral functionalization pattern is described, as well as the first example of an optically pure protected primary amino acid directly linked to the fullerene through only the chiral α-amino-acid carbon atom. An application of an auxiliary chiral nickel-Schiff base moiety as derivatizing agent allowed separation of (f,t A)- and (f,t C)-1,4-fullerene derivatives using an achiral stationary phase, a separation which has never been done before.

Synthesis and biological evaluation of novel 5-hydroxylaminoisoxazole derivatives as lipoxygenase inhibitors and metabolism enhancing agents.

A versatile synthesis of novel 5-hydroxylaminoisoxazoles bearing adamantane moieties has been accomplished using the heterocyclization reactions of readily available unsaturated esters by the treatment with tetranitromethane in the presence of triethylamine and subsequent reduction of resulting 5-nitroisoxazoles by SnCl2 with the participation of THF. A number of obtained isoxazole derivatives were evaluated for their antioxidative activity, inhibition of lipoxygenases and impact on the rat liver mitochondria. The majority of tested compounds demonstrated moderate antiradical activity in DPPH test (up to EC50 16µM). The same compounds strongly inhibited soybean lipoxygenase (up to IC50 0.4µM) and Fe(2+)- and Fe(3+)-induced lipid peroxidation (LP) of rat brain cortex homogenate (up to IC50 0.3µM). All tested isoxazole derivatives promoted the phosphorylating respiratory activity simultaneously with maximal stimulated respiratory activity of mitochondria and do not reveal any toxicity towards the primary culture of rat cortex neurons.