Laboratory of molecular design and synthesis

Department of functioning of living systems

Head: Vladimir Korshun, D.Sc
v-korshun@yandex.ru+7(499)724-67-15

bioconjugation, nucleic acids, fluorescent dyes, Synthetic Methods of Organic Chemistry, Antigens, Gaptens, fluorescent probes, Nonnatural amino acids, Macroheterocycles, chromophores, Ionic liquids, Low molecular bioregulators

  • Obtaining the compounds with antiviral activity.
  • Creating the fluorescent DNA probes.
  • Developing oligonucleotide conjugates as blocks for DNA nanostructures.
  • Creating the mass-spectrometric labels for the detection of biomolecules.
  • Development of innovative methods of synthetic organic chemistry and innovative schemes of synthesis of organic compounds
  • A practical preparation of compounds of different classes, such as benzoheterocycles, arylsulfamides and arylsulfonic acids, polycyclic and cage compounds, arylboronic acids, drug degradation products, antigens and haptens, fluorescent labels, unconventional amino acids, spin-labeled amino acids of the chromophore and photosensitizing compounds, low-molecular weight compounds.
  • The formation of interchain excimer pyrene in the major groove of DNA (ChemBioChem, 2003, 4:841).
  • 1-Phenylethynylpyrene was proposed as improved excimer forming fluorescent label for nucleic acids (Nucleosides Nucleotides, 1997, 16:1461; Eur. J. Org. Chem., 2004, 1298; Bioconjugate Chem., 2007, 18:1972) and used in various types of DNA probes  (Mutation Res., 2006, 599:144; Meth. Mol. Biol., 2009, 578:209; Chem. Eur. J., 2008, 14:11010; Chem. Commun., 2010, 46:8362; Bioconjugate Chem., 2011, 22:533; Analyst, 2016, 141:1331).
  • For the first time, fluorescent dyes were attached to a nucleobase through ethynyl linker (Биоорган. химия, 1996, 22:923; ChemBioChem, 2006, 7:810). A new class of antiviral compounds was found among these nucleosides. The compounds are active against herpes simplex virus type 1, hepatitis C virus and several other enveloped viruses  (Биоорган. химия, 2003, 29:289; Вопросы вирусологии, 2006:34; Tetrahedron, 2006, 62:1279; Org. Biomol. Chem., 2006, 4:1091; PNAS USA, 2010, 107:17339; J. Virol., 2013, 87:3640; Med. Chem. Commun., 2016, 7:495).
  • New mass-spectrometric labels were developed (Org. Biomol. Chem., 2008, 6:4593; Масс-спектрометрия, 2015, 12:253; Analyst, 2016, 141:3289) and the method of single nucleotide polymorphism detection using MALDI spectrometry was proposed (Anal. Chem., 2008, 80:2342; Meth. Mol. Biol., 2009, 578:345) and the analysis method of aminoglycoside antibiotics (Acta Naturae, 2016, 8:128).
  • The application of azide-alkyne cycloaddition was revealed for the synthesis of DNA conjugates (Биоорган. химия, 2010, 36:437) including building blocks for the DNA nanostructures self-assembly (Изв. АН, Сер. хим., 2006:1220; Tetrahedron, 2008, 64:1467; Chem. Commun., 2013, 49:511; Org. Lett., 2014, 16:4590; Tetrahedron, 2016, 72:2386) and for the preparation of efficient fluorogenic DNA probes for real-time PCR assay (Anal. Bioanal. Chem., 2012, 404:59; Analyst, 2014, 139:2867; Analyst, 2016, 141:1331; Anal. Meth., 2016, 8:5826; Mol. Cell. Probes, 2016, 30:285).
  • Oligopeptide components of Plm II protease inhibitors – ω-hydroxy-L-α-amino acids – were synthesized.
  • A new preparative method for the synthesis argiope, synthesized 100 g of this natural toxin. Founded method was used in the synthesis of argiope analogues.
  • Efficient methods for synthesis were designed and representative set of substituted 2-thiazolyl-1-alkyl(aryl)ethanol, prospective templates for new pharmacological agents.
  • Synthetic schemes were developed and synthesis of haptens were conducted, which play an important role in an enzyme immunoassay toxic components of household detergents that pollute the environment, modern pesticides and antibiotics.
  • The methods of synthesis were designed and photoswitchable protein fragment to create a controlled secondary structure elements was received; photoactivated labels for studies of receptor systems; fluorescent labels for the study of structure and functions of nucleic acids; photosensitizing agents; fluorescent indicators.
  • As a part of the Program of the Presidium of the RAS "Biosphere Origin and Evolution" were analyzed the abiogenous variations of the adenine formation conditions and condensation of adenine with ribose and ribosophosphate using the methods of computer synthesis; monosaccharides library were formed, methods of determining the methods of gas chromatography-mass spectrometry and GLC were developed.
  • New subclass of modified nucleosides having high activity against enveloped viruses was founded. Peruse detected earlier (Bioorgan. Chemistry, 29 (3), 289-294 (2003), Nucleosides, Nucleotides & Nucleic Acids, 24 (5/7), 923-926 (2005), Tetrahedron, 62 (6), 1279 -1287 (2006) Problems of Virology, 2006 (1), 34-38, Org. Biomol. Chem., 4 (6), 1091-1096 (2006)) identified a class of antiviral compounds from the group of substances active against enveloped viruses. Specifically, IC50 5- (perylene-3-yl) ethynyl-2'-deoxyuridine was 50 nmol / L for herpes simplex virus type 1 and 180 nmol / L for hepatitis C. A characteristic feature of active nucleosides structure is the availability of an aromatic residue hydrocarbon perylene rigidly attached to the nucleobase via a triple bond. It is suggested that the unique structure of the resulting compounds is responsible for their incorporation into the virion membrane, resulting in the fusion of the virion to the cell (infection) becomes very disadvantageous process (Figure). The novel compounds practically cytotoxic.

All publications (show selected)

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Vladimir Korshun

Antiviral compounds based on 5-(perylen-3-ylethynyl)uracil scaffold

In collaboration with Group of Cross-Linking Enzymes

Rigid amphipathic fusion inhibitors (RAFIs) are potent antivirals based on a perylene core linked with a nucleoside moiety. Sugar-free analogues of RAFIs, 5-(perylen-3-ylethynyl)uracil-1-acetic acid and its amides were synthesized using combined protection group strategy. Compounds appeared to have low toxicity on porcine embryo kidney (PEK) or rhabdomiosarcoma (RD) cells together with remarkable activity against enveloped tick-borne encephalitis virus (TBEV): EC50 values vary from 0.077 μM to subnanomolar range. Surprisingly, 3-pivaloyloxymethyl (Pom) protected precursors showed even more pronounced activity. The propargylamide of N3-Pom-protected 5-(perylen-3-ylethynyl)uracil acetic acid, a universal precursor, was used in CuAAC click reaction for the synthesis of several derivatives including three ramified molecules with high activities against tick borne encephalitis virus (TBEV). Pentaerythritol-based polyazides were used for the assembly of molecules containing 2…4 antiviral 5-(perylen-3-ylethynyl)uracil scaffolds, the first examples of polyvalent perylene antivirals. Four prepared compounds, including one ramified cluster, showed remarkable 1…3 nM EC50 values against TBEV in cell culture. The results suggest that the carbohydrate moiety of RAFI nucleosides does not play a crucial role in their antiviral action, and biological activity of the 5-(perylen-3-ylethynyl)uracil scaffold can be effectively modulated by substituents in positions 1 and 3. The high antiviral activity of new compounds, coupled with low toxicity advocate their potential role in antiviral therapy. The antiviral evaluations were performed in Chumakov Institute of Poliomyelitis and Viral Encephalitides.

Fluorescent Yin-Yang probes for RT-qPCR detection of low copy HIV RNA

Nucleic acids labeled with a fluorophore/quencher pair are widely used as probes in biomedical research and molecular diagnostics. We synthesized novel DNA molecular beacons double labeled with the identical dyes (R6G, ROX and Cy5) at 5′- and 3′-end, studied their photophysical properties, and demonstrated that fluorescence quenching by formation of the homo dimer exciton in such molecular beacons allows using them in homogeneous assays. Further, we developed and evaluated homo Yin-Yang DNA probes labeled with identical dyes and used them for detection of low copy HIV RNA by RT-qPCR. They demonstrated improved sensitivity (LLQ: 10 vs 30 copies mL-1) in comparison to commercially available Abbott RealTime HIV-1 kit based on VIC-BHQ dyes both for model mixtures (naive human plasma with added deactivated HIV-1 virus) and for preliminarily confirmed 36 clinical samples (4 vs 1 positive ones for low-copy samples). The research was performed in collaboration with ScolTech, Research Institute of Epidemiology, and other institutions.

Publications

  1. (book) Kireev DE, Farzan VM, Shipulin GA, Korshun VA, Zatsepin TS (2020). RT-qPCR Detection of Low-Copy HIV RNA with Yin-Yang Probes. Methods Mol Biol 2063, 27–35
  2. Farzan VM, Kvach MV, Aparin IO, Kireev DE, Prikazchikova TA, Ustinov AV, Shmanai VV, Shipulin GA, Korshun VA, Zatsepin TS (2019). Novel homo Yin-Yang probes improve sensitivity in RT-qPCR detection of low copy HIV RNA. Talanta 194, 226–232

The structure of the two components of the lipopeptide antibiotic crystallomycin from a sample obtained 60 years ago has been established. The identity of the components of two crystallomycin components to these of aspartocin (the structure of which has been elucidated recently) has been found. The antibiotic exhibits Ca2+ -dependent activity against gram-positive bacteria. The conformations of crystallomycin 2 in solution were investigated using NMR.

The amino acid 4-chloro-L-kinurenin, previously found in natural products only once, was found in the peptide antibiotic INA-5812. We first described the fluorescent properties of 4-chloro-L-kinurenin and its use as an energy donor for the excitation of other fluorophores.

The structure of two new macrolide antibiotics, astolides A and B, has been established using various 2D NMR techniques. Astolide molecules contain simultaneously a membrane-active polyol macrolide and a redox-active naphthoquinone residue as aglycones. The presence of a hydroxyl group at position 18 dramatically changes the spectrum of biological activity in comparison with the known analogues – antifungal activity increases and cytotoxicity reduces.

Publications

  1. Alferova VA, Shuvalov MV, Suchkova TA, Proskurin GV, Aparin IO, Rogozhin EA, Novikov RA, Solyev PN, Chistov AA, Ustinov AV, Tyurin AP, Korshun VA (2018). 4-Chloro-l-kynurenine as fluorescent amino acid in natural peptides. Amino Acids 50 (12), 1697–1705
  2. Alferova VA, Novikov RA, Bychkova OP, Rogozhin EA, Shuvalov MV, Prokhorenko IA, Sadykova VS, Kulko AB, Dezhenkova LG, Stepashkina EA, Efremov MA, Sineva ON, Kudryakova GK, Peregudov AS, Solyev PN, Tkachev YV, Fedorova GB, Terekhova LP, Tyurin AP, Trenin AS, Korshun VA (2018). Astolides A and B, antifungal and cytotoxic naphthoquinone-derived polyol macrolactones from Streptomyces hygroscopicus. Tetrahedron 74 (52), 7442–7449
  3. Jiang ZK, Tuo L, Huang DL, Osterman IA, Tyurin AP, Liu SW, Lukyanov DA, Sergiev PV, Dontsova OA, Korshun VA, Li FN, Sun CH (2018). Diversity, novelty, and antimicrobial activity of endophytic actinobacteria from mangrove plants in Beilun Estuary National Nature Reserve of Guangxi, China. Front Microbiol 9 (MAY), 868
  4. Tyurin AP, Alferova VA, Paramonov AS, Shuvalov MV, Malanicheva IA, Grammatikova NE, Solyev PN, Liu S, Sun C, Prokhorenko IA, Efimenko TA, Terekhova LP, Efremenkova OV, Shenkarev ZO, Korshun VA (2018). Crystallomycin revisited after 60 years: Aspartocins B and C. Medchemcomm 9 (4), 667–675

New antiviral nucleoside derivatives for inhibiting the reproduction of varicella-zoster virus and tick-borne encephalitis virus

In collaboration with The group of molecular tools for living system studies

A series of analogues of potent antiviral perylene nucleoside dUY11 with methylthiomethyl (MTM), azidomethyl (AZM) and HO-C1-4-alkyl-1,2,3-triazol-1,4-diyl groups at 3`-O-position as well as the two products of copper-free alkyne-azide cycloaddition of the AZM derivative were prepared and evaluated against tick-borne encephalitis virus (TBEV). Four compounds showed EC50 ≤10 nM, thus appearing the most potent TBEV inhibitors to date. Moreover, these nucleosides have higher lipophilicity (clogP) and increased solubility in aq. DMSO vs. parent compound dUY11.

Phenoxazine scaffold is widely used to stabilize nucleic acid duplexes, as a part of fluorescent probes for the study of nucleic acid structure, recognition, and metabolism etc. We present the synthesis of phenoxazine-based nucleoside derivatives and their antiviral activity against a panel of structurally diverse viruses: enveloped DNA herpesviruses varicella zoster virus (VZV) and human cytomegalovirus, enveloped RNA tick-borne encephalitis virus (TBEV), and non-enveloped RNA enteroviruses. Studied compounds were effective against DNA and RNA viruses reproduction in cell culture. 3-(2’-Deoxy-β-D-ribofuranosyl)-1,3-diaza-2-oxophenoxazine proved to be a potent inhibitor of VZV replication with superior activity against wild type than thymidine kinase deficient strains (EC50 0.06 and 10 µM, respectively). This compound did not show cytotoxicity on all the studied cell lines. Several compounds showed promising activity against TBEV (EC50 0.35-0.91 µM), but the activity was accompanied with pronounced cytotoxicity. These compounds may be considered as a good starting point for further structure optimization as antiherpesviral or antiflaviviral compounds.

Publications

  1. Proskurin GV, Orlov AA, Brylev VA, Kozlovskaya LI, Chistov AA, Karganova GG, Palyulin VA, Osolodkin DI, Korshun VA, Aralov AV (2018). 3′-O-Substituted 5-(perylen-3-ylethynyl)-2′-deoxyuridines as tick-borne encephalitis virus reproduction inhibitors. Eur J Med Chem 155, 77–83