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Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia
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research activity
  • oligosaccharide synthesis
  • neoglycoconjugates
  • glycoarrays
  • supramolecular chemistry
  • influenza
  • transplantology and blood groups
  • oncology
  • natural antibodies
  • glycotargeting
  • The laboratory focussed at basic problems of glycobiology and application of glyco-molecules in medicine. The main directions comprise synthesis of oligosaccharides and glycoconjugates, self-assembling peptides, carbohydrate-protein interactions, innate immunity to carbohydrates, study of cell surface lectins, influenza virus. Our current glyco-medical project are:
    Neoglycoconjugates. click to enlarge...

    We have developed a versatile approach to the synthesis of neoglycoconjugates (see review). The method is based on attachment of carbohydrate and non-carbohydrate ligands (labels, effectors) to activated polymer. The glycoconjugates are useful for flow cytometry, fluorescent microscopy, ELISA, beads modification, etc.
    Glycoarrays. click to enlarge...

    Our lab has developed a glycan microarray (printed glycan array, PGA) in cooperation with the Consortium for Functional Glycomics. Glycoarray is a glass or plastic slide 25 × 75 mm (1 × 3 inches). Ligands are immobilized on its surface as the spots having ~100 μm diameter. Its plastic version is under development by Semiotik LLC as key platform for cancer diagnostics and reproduction deseases.
    Current version includes ~400 synthetic oligosaccharides and ~200 bacterial polysaccharides.

    Ligands on microarray
    Applications:
    Supramolecular chemistry: synthesis of self-assembling peptides and glycopeptides. click to enlarge...

    The interest to small molecule self-assembling is caused to high extent by the possibility to design nanomaterials with customized properties and molecular devices. The driving force of self-assembling is simultaneous formation of multiple hydrogen bonds between complementary fragments. Our goal was the design of non-covalent polymers, (Glycosyl-Z)n, where Z fragment provides the assembling, soluble and stable in aqueous solution. We expected that the supramers could display sharply increased activity due to multivalent display of Glycosyl functionality When synthesizing oligovalent glycopeptides, [glycinen-NHCH2]4C, we have observed that some of them possess the required properties, i.e. form supramers that are stable in aqueous solutions. Oligopeptide chains (glycosyl free) are joined in the supramers by polyglycine II type, forming water insoluble flat sheets of submicron size, one molecule thick (~50Å). Reversible disassembling takes place in concentrated solution of lithium bromide, trifluoroacetic acid, or upon heating. Due to rigidity, regulated thickness, NH2functionality, and reversibility of self-assembly obtained material is prospective as atomically smooth platform for nano-device engineering.

    Inhibitors of influenza virus adhesion. click to enlarge...

    The aim of this study is the revealing of mechanism and topography of influenza virus-to-cell first contact, and, finally, the development of antiviral preparation acting by principle of inhibitionof the adhesion. Sels-assembling glyco-glycines (see Supramolecular chemistry) work well as anti-adhesion therapeutics on ferret model.

    Transplantology and blood groups. click to enlarge...

  • Interaction of natural blood group anti-A and anti-B antibodies of recipient with the corresponding antigens of transplanted material often hinders or makes difficult allotransplantation of organs and tissues because antigens A and B are present not only on erythrocytes but on many other organs. A practically convenient way to overcome this obstacle is affinity removal of the corresponding antibodies. We have developed adsorbents on the base of SepharoseFF™ and synthetic glycans A and B attached to the matrix via flexible hydrophilic polymer. Company Pocard (Moscow) produces commercial affinity A ans B columns for the antibodies removal.
  • Lewis serologic reagents frequently give inaccurate phenotyping results. Knowledge of their fine-specificity and potential cross-reactivity will improve the quality of the assaya. A range of Lewis reagents including mouse monoclonal (MAb), human and goat polyclonal (PAb) reagents were evaluated. All were evaluated against both Lewis kodecytes expressing only single Lea, Leb, ALeb, BLeb, Lex, Ley, ALey or BLey antigens and against the same antigens inkjet printed on a paper-based microplate and analysed by enzyme immunoassay. Nine clinical samples were also evaluated. A kodecyte (kodecytes are human RDC with incerted synthetic glycolipids) antigen-dilution sensitivity assay was used to establish the ratio of Leb antigen between group A1/A2 and O RBCs.
  • A continuum of cross-reactivity from Lex through to H was observed with MAbs. All PAb and few MAb anti-Lea samples and reagents cross-reacted to some degree with Leb antigen. Some PAb and MAb anti-Leb did not cross-react with Lea. All polyclonal goat anti-Leb reagents showed substantial activity against ALeb and BLeb, while no MAb reagent had this activity. A1 RBCs had less than half the Leb antigen of A2/O RBCs. Thus, substantial cross-reactivity of both MAbs and PAbs with related antigens highlights the risks of using serologic reagents in non-serologic assays or against synthetic antigens. The lack of ALeb activity in anti-Leb MAbs explains their poor performance against blood group A1 Le(a-b+) phenotypes.
    Although the nature of the blood groups A and B has been comprehensively studied for a long time, it is still unclear as to what exactly is the epitope that is recognized by antibodies having AB specificity, i.e. monoclonal and polyclonal antibodies which are capable of interacting equally well with the antigens GalNAcα1-3(Fucα1-2)Gal (A trisaccharide) and Galα1-3(Fucα1-2)Gal (B trisaccharide), but do not react with their common fragment Fucα1-2Gal. We have supposed that besides Fucα1-2Gal, A and B antigens have one more shared epitope. The trisaccharides A and B are practically identical from the conformational point of view, the only difference being situated at position 2 of Galα residue, i.e. trisaccharide A has a NHAc group, whereas trisaccharide B has a hydroxyl group. We have hypothesized that the AB-epitope should be situated in the part of the molecule that is opposite to the NHAc group of GalNAc residue. In order to test this hypothesis we have synthesized a polymeric conjugate in such a way that de-N-acetylated A-trisaccharide is attached to a polymer via the nitrogen in position C-2 of the galactosamine residue. In this conjugate the supposed AB-epitope should be maximally accessible for antibodies from the solution, whereas the discrimination site of antigens A and B by the antibodies should be maximally hidden due to the close proximity of the polymer. Interaction with several anti-AB monoclonal antibodies revealed that a part of them really interacted with the synthetic AB-glycotope, thus confirming our hypothesis. Moreover, similar antibodies were revealed in the blood of healthy blood group 0 donors. Analysis of spatial models was performed in addition to identify the hydroxyl groups of Fuc, Galα, and Galβ residues, which are particularly involved in the composition of the AB-glycotope.

    Oncology. click to enlarge...

  • Antibodies to Galβ1-3GlcNAc (LeC) disaccharide found to have high titers in practically all healthy donors. In breast cancer patients its level is significantly reduced, therefore we supposed that the antibody play a role of housekeepers for elimination of cancer cells. Isolated monospecific anti-LeC specifically stained breas cancer cells. In 2017 we started project directed to application of the antibodies as anti-cancer therapeutic.
  • Printed glycan array allows us to find in blood of cancer patients diagnostically relevant anti-glycan natural antibodies (diagnostic signature) for revealing of breast, ovarian and colorectal cancers.
  • Supposing that a tumor is an inflammation site, selectin ligand SiaLex was used as specific vector (lipophilic form of the tetrasaccharide) for delivery of cytotoxic liposomes to tumor. Indeed, such liposomes caused drastic increase in life time of mice with grafted tumors, even in case of single preparation administration.
  • The role of carbohydrate chains in phagocytosis of apoptotic bodies obtained from melanoma cell line has been studied on a model where macrophage cell line was used. It was shown that termination Galβ1-3GalNAcβ is the specific component of apoptotic bodies surface during phagocytosis, whereas the specific component of phagocyting cells is, possibly, galectin-1. Moreover, synthetic lipophilic variant of the disaccharide built in apoptotic bodies and administered to experimental mice caused phagocytosis increase and elevated vitality of animals with experimental tumors.

    Natural antibodies. click to enlarge...

    It is generally accepted that the generation of antibodies proceeds due to immunization of an organism by alien antigens, and the level and affinity of antibodies are directly correlated to the presence of immunogen. At the same time, mumerous experimental data provide evidence of antibodies whose level remains unchanged and affinity is constant during a lifetime. In contrast to the first, adaptive immunoglobulins, the latter are named natural antibodies (nAbs). The nAbs are produced by B1 cells, whereas adaptive Abs are produced by B2. Using printed glycan array (PGA) we identified big number of nAbs not described earlier, including cancer-associated anti-LeC. We use hapten-specific affinity chromatography for isolation of human and mammalian nAbs with subsecuent PGA analysis for investigation of their fine epitope specificity. Using mice model we study genesis of nAbs and role of microbiota in the genesis and regulation of nAbs.