Triterpenoid pyrazines and pyridines - Synthesis, cytotoxicity, mechanism of action, preparation of prodrugs.

A set of fifteen triterpenoid pyrazines and pyridines was prepared from parent triterpenoid 3-oxoderivatives (betulonic acid, dihydrobetulonic acid, oleanonic acid, moronic acid, ursonic acid, heterobetulonic acid, and allobetulone). Cytotoxicity of all compounds was tested in eight cancer and two non-cancer cell lines. Evaluation of the structure-activity relationships revealed that the triterpenoid core determined whether the final molecule is active or not, while the heterocycle is able to increase the activity and modulate the specificity. Five compounds (1b, 1c, 2b, 2c, and 8) were found to be preferentially and highly cytotoxic (IC50 ≈ 1 µM) against leukemic cancer cell lines (CCRF-CEM, K562, CEM-DNR, or K562-TAX). Surprisingly, compounds 1c, 2b, and 2c are 10-fold more active in multidrug-resistant leukemia cells (CEM-DNR and K562-TAX) than in their non-resistant analogs (CCRF-CEM and K562). Pharmacological parameters were measured for the most promising candidates and two types of prodrugs were synthesized: 1) Sugar-containing conjugates, most of which had improved cell penetration and retained high cytotoxicity in the CCRF-CEM cell line, unfortunately, they lost the selectivity against resistant cells. 2) Medoxomil derivatives, among which compounds 26-28 gained activities of IC50 0.026-0.043 µM against K562 cells. Compounds 1b, 8, 21, 22, 23, and 24 were selected for the evaluation of the mechanism of action based on their highest cytotoxicity against CCRF-CEM cell line. Several experiments showed that the majority of them cause apoptosis via the mitochondrial pathway. Compounds 1b, 8, and 21 inhibit growth and disintegrate spheroid cultures of HCT116 and HeLa cells, which would be important for the treatment of solid tumors. In summary, compounds 1b, 1c, 2b, 2c, 24, and 26-28 are highly and selectively cytotoxic against cancer cell lines and were selected for future in vivo tests and further development of anticancer drugs.

Substituted dienes prepared from betulinic acid - Synthesis, cytotoxicity, mechanism of action, and pharmacological parameters.

A set of new substituted dienes were synthesized from betulinic acid by its oxidation to 30-oxobetulinic acid followed by the Wittig reaction. Cytotoxicity of all compounds was tested in vitro in eight cancer cell lines and two noncancer fibroblasts. Almost all dienes were more cytotoxic than betulinic acid. Compounds 4.22, 4.30, 4.33, 4.39 had IC50 below 5 µmol/L; 4.22 and 4.39 were selected for studies of the mechanism of action. Cell cycle analysis revealed an increase in the number of apoptotic cells at 5 × IC50 concentration, where activation of irreversible changes leading to cell death can be expected. Both 4.22 and 4.39 led to the accumulation of cells in the G0/G1 phase with partial inhibition of DNA/RNA synthesis at 1 × IC50 and almost complete inhibition at 5 × IC50. Interestingly, compound 4.39 at 5 × IC50 caused the accumulation of cells in the S phase. Higher concentrations of tested drugs probably inhibit more off-targets than lower concentrations. Mechanisms disrupting cellular metabolism can induce the accumulation of cells in the S phase. Both compounds 4.22 and 4.39 trigger selective apoptosis in cancer cells via intrinsic pathway, which we have demonstrated by changes in the expression of the crucial apoptosis-related protein. Pharmacological parameters of derivative 4.22 were superior to 4.39, therefore 4.22 was the finally selected candidate for the development of anticancer drug.

Axially Chiral Trifluoromethylbenzimidazolylbenzoic Acid: A Chiral Derivatizing Agent for α-Chiral Primary Amines and Secondary Alcohols To Determine the Absolute Configuration.

Racemic 2-(2-trifluoromethyl)-1H-benzo[d]imidazol-1-yl)benzoic acid (TBBA) was synthesized in three steps from 1-fluoro-2-nitrobenzene. Target (P)- and (M)-TBBA atropisomers were stable with a racemization barrier above 30 kcal/mol. As a chiral derivatizing agent, TBBA showed much higher differences in chemical shifts (ΔδPM) than the conventional Mosher's acid.

Intralaboratory comparison of analytical methods for quantification of major phytocannabinoids.

This study compares alternative approaches for analyzing phytocannabinoids in different plant materials. Three chromatographic analytical methods (ultra-high-performance liquid chromatography with tandem mass spectrometric detection and gas chromatography with mass spectrometric and flame ionization detection) were evaluated regarding selectivity, sensitivity, analytical accuracy, and precision. The performance of the methods was compared and all three methods were demonstrated to be appropriate tools for analyzing phytocannabinoids in cannabis. Gas chromatography coupled with mass spectrometric detection showed slightly better accuracy in determining phytocannabinoid acids, which are often difficult to quantify owing to their limited stability. Aspects of sample preparation, such as material homogenization and extraction, were also considered. A single ultrasonic-assisted ethanolic extraction of dried and powdered plant samples of cannabis was shown to be exhaustive for extracting the samples prior to analysis.

Polymer-Assisted Synthesis of Single and Fused Diketomorpholines.

The synthesis of different diketomorpholines via N-acyl-3,4-dihydro-2 H-1,4-oxazine-3-carboxylic acids is reported in this article. The key intermediates were prepared using a convenient solid-phase synthesis starting from polymer-supported Ser( tBu)-OH. After subsequent sulfonylation with 4-nitrobenzenesulfonyl chloride (4-Nos-Cl), alkylation with an α-bromoketone, cleavage of the 4-Nos group and acylation with an α-halocarboxylic acids, acid-mediated cleavage from the resin yielded dihydrooxazine-3-carboxylic acids in high crude purities. Depending on the reaction conditions, exposure to base resulted in cyclization to either oxazino[3,4- c][1,4]oxazine-diones or 3-methylidenemorpholine-2,5-diones. Further reaction with triethylsilane-trifluoroacetic acid (TES/TFA) led to olefin reduction, in the case of oxazino[3,4- c][1,4]oxazine-dione with full control of the newly formed stereocenter.

A novel three-fluorophore system as a ratiometric sensor for multiple protease detection.

A synthetic three-fluorophore system with two enzymatically cleavable linkers has been developed for the simultaneous detection of two proteases in a mixture. The probe was designed to afford single excitation/triple emission ratiometric detection through a fluorescence change during the cleavage of a peptide linker. The developed assays were verified for trypsin and chymotrypsin as the model enzymes.