Andrey G. Pokrovsky

Discovery of a new class of antiviral compounds: camphor imine derivatives.

A new class of compounds featuring a camphor moiety has been discovered that exhibits potent inhibitory activity against influenza A(H1N1)pdm09 and A(H5N1) viruses. The synthesized compounds were characterized by spectroscopic analysis; in addition the structures of compound 2 and 14 were elucidated by the X-ray diffraction technique. Structure-activity relationship studies have been conducted to identify the 1,7,7-trimethylbicyclo[2.2.1]heptanes2-ylidene group as the key functional group responsible for the observed antiviral activity. The most potent antiviral compound is imine 2 with therapeutic index more than 500.

Structural, biocomplexation and gene delivery properties of hydroxyethylated gemini surfactants with varied spacer length.

Gemini surfactants with hexadecyl tails and hydroxyethylated head groups bridged with tetramethylene (G4), hexamethylene (G6) and dodecamethylene (G12) spacers were shown to self-assemble at the lower critical micelle concentration compared to their conventional m-s-m analogs. The lipoplex formation and the plasmid DNA transfer into different kinds of host cells were studied. In the case of eukaryotic cells, high transfection efficacy has been demonstrated for DNA-gemini complexes, which increased as follows: G6<G4<G12. Different activity series, i.e., G6>G4>G12 has been obtained in the case of transformation of bacterial cells with plasmid DNA-gemini complexes, mediated by electroporation technique. Solely G6 shows transformation efficacy exceeding the control result (uncomplexed DNA), while the inhibitory effect occurs for G4 and G12. Analysis of physico-chemical features of single surfactants and lipoplexes shows that compaction and condensation effects change as follows: G6<G4 ≤ G12, i.e., agree with the order of transfection efficacy, which is supported by membrane tropic properties of G12. On the other hand, gel retardation assay and docking study testify low electrostatic affinity in G12/DNA pair, thereby indicating that hydrophobic effect probably plays important role in the lipoplex formation. Two factors are assumed to be responsible for the inhibition effect of gemini in the case of transformation of bacterial cells. They are (i) an unfavorable influence of cationic surfactants on the electroporation procedure due to depressing the electrophoretic effect; and (ii) antibacterial activity of cationic surfactants that may cause the disruption of integrity of cell membranes.

Synthesis of 1H-1,2,3-triazole linked aryl(arylamidomethyl) - dihydrofurocoumarin hybrids and analysis of their cytotoxicity.

A series of 2-(4-R-triazolyl)substituted 3-oxo-2,3-dihydrofurocoumarins have been synthesized by a regioselective cycloaddition of 2-azidooreoselone 1 or 2-azido-9-[(4-methylpiperazin-1-yl)methyl]oreoselone 2 with various alkynes in the presence of Cu(II)/ascorbate in water/methylene chloride reaction medium. The structure of 2-azidooreoselone was established by X-ray structure analysis. The cytotoxicity of 2-substituted dihydrofurocoumarins was determined against three cancer cell lines (CEM-13, MT-4, U-937) using the conventional MTT assays. Among the tested molecules, most of the analogs displayed better cytotoxic activity then the parent natural furocoumarin peucedanin 3. The activity and selectivity to the cell line increased even further in the series of 2-(4-{2,3-dihydrobenzo[b][1,4]dioxine}triazolyl)-3-oxo-2,3-dihydrofurocoumarins and 2-(4-aryltriazolyl)-3-oxo-2,3-dihydrofurocoumarins having the (4-methylpiperazin-1-ylmethyl) substituent in the 9-th position. The most active compound 20 contain the 4-hydroxy-3-methoxybenzamidomethyl substituent in the 4-th position at the triazole ring of 2-(triazol-1-yl)dihydrofurocoumarins. The obtained 2-triazolyl substituted dihydrofurocoumarins were studied as inhibitors of phosphodiesterase (PDE-4B) using docking experiments. As a result of virtual screening 3 compounds are selected based on minimum binding energy. The interactions of the most active compound and amino acid residues in the binding site were studied.

Cationic gemini surfactants as new agents for plasmid DNA delivery into cells

197 Gene therapy has a great potential for treating many human diseases that are currently considered incurable. Viral vectors based on adenoviruses or rett roviruses are very effective in gene delivery. Neverthe less, in view of the problems associated with immunoo genicity and biosafety of viral vectors, there remains a need to develop nonviral vectors. As an alternative to viral vectors, cationic polymers [1–3] and liposomes [4], which bind to DNA through electrostatic interacc tions and form nanosized complexes, have been invess tigated [5, 6]. Cationic agents protect DNA from degg radation by nucleases and serve as mediators in the penetration into the cell and subsequent release from endosomes, which increases the efficiency of transfecc tion. The use of cationic surfactants as nonviral vectors was described in several papers [6, 7]. It is shown that they are effective agents that provide the compaction of DNA and recharging of the complex. Despite the obvious advantages (simplicity of synthesis and forr mulation, low concentration, availability, and high complexing ability), a significant drawback of synn thetic agents based on cationic surfactants is the low transfection efficiency. In view of this, the search for new cationic vector systems is a relevant problem. In this study, we investigated alkylammonium gemm ine surfactants (AGSs) with the formula , where m = 4–12, R = СН 3 , С 2 Н 4 ОН, R 1 = nC n Н 2n + 1 , where n = 10, 12, 14, and 16. Aqueous solutions of these compounds ensure efficient delivery of DNA into cells. Gemine surfactants (alkanedyllα,ω bis(methyldialkylammonium bromides)) were pree pared under laboratory conditions as described in [8]. The structure of the obtained compounds was conn firmed by elemental analysis as well as by IR and NMR spectroscopy data. Cell line 293T (human embryonic kidney epithee lium cells), which was used as a producer of pseudoo lentiviruses and as a model target cell line, was mainn tained by the standard procedure. We used the peGFPPN1 plasmid (Clontech, United States), which is 5100 bp in length and expresses the green fluoress cent protein, and the pCIINEO plasmiddbased vector (Promega, United States), which is 5000 bp in length and does not contain the green fluorescent protein gene. HEK293T cells were replated 40 thousand cells per well one day before transfection. To transfect HEK293T cells, 50–100 thousand cells per well of a 966well plate were used. One hour before transfection, …

Synthesis and cytotoxic activity of a new group of heterocyclic analogues of the combretastatins.

A series of new analogs of combretastatin A-4 (CA-4, 1) with the A or B-ring replaced by a 3-oxo-2,3-dihydrofurocoumarin or a furocoumarin residue have been designed and synthesized by employing a cross-coupling approach. All the compounds were evaluated for their cytotoxic activity with respect to model cancer cell lines (CEM-13, MT-4, U-937) using conventional MTT assays. Structure-activity relationship analysis reveals that compounds 2, 3, 6–8 in which the (Z)-styryl substituent was connected to the 2-position of the 3-oxo-2,3-dihydrofurocoumarin core, demonstrated increased potency compared to 3-(Z)-styrylfurocoumarins 4, 5, 9–11. The methoxy-, hydroxyl- and formyl- substitution on the aromatic ring of the (Z)-styryl moiety seems to play an important role in this class of compounds. Compounds 2 and 3 showed the best potency against the CEM-13 cell lines, with CTD50 values ranging from 4.9 to 5.1 μM. In comparison with CA-4, all synthesized compounds presented moderate cytotoxic activity to the T-cellular human leucosis cells MT-4 and lymphoblastoid leukemia cells CEM-13, but most of them were active in the human monocyte cell lines U-937.

Phenolic compounds from Glycyrrhiza pallidiflora Maxim. and their cytotoxic activity

Abstract Twenty-one phenolic compounds (1–21) including dihydrocinnamic acid, isoflavonoids, flavonoids, coumestans, pterocarpans, chalcones, isoflavan and isoflaven, were isolated from the roots of Glycyrrhiza pallidiflora Maxim. Phloretinic acid (1), chrysin (6), 9-methoxycoumestan (8), isoglycyrol (9), 6″-O-acetylanonin (19) and 6″-O-acetylwistin (21) were isolated from G. pallidiflora for the first time. Isoflavonoid acetylglycosides 19, 21 might be artefacts that could be produced during the EtOAc fractionation process of whole extract. Compounds 2–4, 10, 11, 19 and 21 were evaluated for their cytotoxic activity with respect to model cancer cell lines (CEM-13, MT-4, U-937) using the conventional MTT assays. Isoflavonoid calycosin (4) showed the best potency against human T-cell leukaemia cells MT-4 (CTD50, 2.9 μM). Pterocarpans medicarpin (10) and homopterocarpin (11) exhibit anticancer activity in micromolar range with selectivity on the human monocyte cells U-937. The isoflavan (3R)-vestitol (16) was highly selective on the lymphoblastoid leukaemia cells CEM-13 and was more active than the drug doxorubicin

The short way to chiral compounds with hexahydrofluoreno(9,1-bc) furan framework: Synthesis and cytotoxic activity

A simple and efficient method for synthesizing chiral heterocyclic compounds with the hexahydrofluoreno[9,1-bc]furan framework via interaction between trans-4-hydroxymethyl-2-carene and aromatic aldehydes containing methoxy and hydroxyl moieties in the presence of montmorillonite clay was found. One of the synthesized compounds exhibited a high cytotoxic activity against lymphoblastoid cell line MT-4 (CTD50 0.9μM), which was higher than that of the comparative drug Doxorubicin. Death of cancer cells in this case substantially occurs via induction of apoptosis.

Cytotoxic and Antiviral Properties of Novel Dehydroabietylamine Salts

Here, we report the synthesis of a series of dehydroabietylamine (DAA) salts. We studied the cytotoxic activity of DAA salts and their and antiviral properties against influenza virus A(H1N1)pdm09 We further compared these parameters to those of DAA itself and to organic acids used as counterions. We observed that the DAA, its hydrochloride (DAA∙HCl) and organic salts had similar cytotoxicity profiles in three cancer cell lines: MDA-MB-231, MCF-7, U-87 MG. Meanwhile the cytotoxicity of same organic salts of DAA with respect to melanoma cell line SK-Mel-8 was higher than those DAA and DAA∙HCl: CTD50 values of 5 from 14 synthesized salts were 2-2.8 times lower than that of CTD50 for DAA∙HCl. Pure organic acids used for the synthesis of DAA salts were noncytotoxic to cancer cells, except ursolic and glycyrrhetic acid having moderate cytotoxicity. Further, we found that modification of DAA with counterions derived from the selected organic acids does not affect, in most cases, the DAA antiviral activity (except compound 4n, DAA glycyrrhetate). The toxicity of this compound was significantly lower than that of its constituting ions, thereby making the selectivity index of this compound against the A(H1N1) pdm09 virus several times higher than that of other DAA salts.

Development and validation of ultrafast LC–MS/MS method for quantification of anti-influenza agent camphecene in whole rat blood using dried blood spots and its application to pharmacokinetic studies

A fast, selective and sensitive procedure for quantitation of the camphor-based anti-influenza agent camphecene in whole rat blood was developed and validated using dried blood spots and LC-MS/MS. The method was validated according to recommendations of the FDA and EMA in terms of selectivity, linearity, accuracy, precision, recovery, matrix factor, stability, and carry-over. Sample preparation included spotting 20μL of whole blood taken from the tail vein onto the paper, drying and extracting the analyte, followed by evaporation of the solvent and analysis of the residue. HPLC separations were run on a reversed-phase microcolumn; the time of analysis was less than 2min. MS/MS detection was performed on a triple quadrupole mass-spectrometer using multiple reaction monitoring (MRM) mode. Transitions 196.4→122.2/153.3 and 152.2→93.1/107.2 were monitored for camphecene and 2-adamantylamine hydrochloride (internal standard), respectively. The intra- and inter-day precisions and accuracies, matrix factor, carry-over and recovery were within acceptable limits. Despite low extraction recovery (less than 2%), the sensitivity of the method was enough to detect the analyte in the concentration range 50-2500ng/mL. The application of the method was shown in pharmacokinetic studies of camphecene in rats at a dose of 10mg/kg.

Lipoplexes of dicationic gemini surfactants with DNA: Structural features of DNA compaction and transfection efficiency

The internal structure of DNA lipoplexes with hydroxyethylated alkylammonium gemini surfactants (GS) with high transfection activity was studied by circular dichroism. It was shown that the efficiency of transfection of HEK293T cells with the pEGFP-N1 circular plasmid was different from zero only in the region of existence of chiral supramolecular DNA-GS complexes and reaches a maximum at concentrations at which the spontaneous aggregation of components is observed.