Organic Synthesis and NMR Spectroscopy SRL

Address: 85 Pobedy Street, Building 10, Office 1-11 Phone: +7 4722 301327 Head of the Laboratory – Anatoliy Buchelnikov, Candidate of Mathematics Sciences

• The SRL basic activities and researches in the synthesis of pharmaceutical substances, biologically active compounds and colorants
• The main activities and laboratory projects in the research field of complex formation of biologically active compound with bioreceptors
• Other activities of the laboratory

Organic Synthesis and NMR Spectroscopy SRL was founded in September 2013 as biomolecule NMR Spectroscopy SRL. Since June 2015 due to the coming of Boris Paponov, Candidate of Сhemical Sciences, range of SRL activities includes the researches in innovative pharmaceutical substances synthesis.

The SRL basic activities and researches in the synthesis of pharmaceutical substances, biologically active compounds and colorants

Head of research area – senior fellow Boris Paponov.

The SRL main research area is engineering, synthesis and identification of synthetic compounds and modification of natural compounds with targeted properties. First of all this is the search of compounds with targeted biological activities and colorants with targeted spectral-luminescent properties ( including cells colorants).

The SRL solves the following scientific and applied scientific problems:

The main scientific aim is to process original low-molecular ligands for biotarget, including protein inhibitor kinase for treatment of oncological, neurodegenerative, inflammatory, and autoimmune diseases. The central focus is on the compounds effecting on proliferation, differentiation and transformation of cells, including different types of stem cells, in particular cancer stem cells. Laboratory personnel deals with high-selective and multi-tyrosine kinase inhibitor. The target – ATP-binding activity.

An interesting aspect of this research area is the creation of synthetical mimetics of such natural biologically active substances as indirubins, meriolins, ariolins etc.

Another target is imidazoline receptors. Here the main aim is engineering and synthesis of low-molecular compounds as efficient antidiabetic drugs for type 2 diabetes.

To solve following problems we apply for engineering:

• Application of bioinformatics and gene regulatory cell cycle network methods for identification of new biotargets and their combination in order to create innovative biologically active compounds and find optimal combination of the permitted for use drugs with new activities.
• Task-oriented screening in data basis and knowledge base with the help of modern bioinformatics methods of pharmacologic compounds and their combinations which were tested in vitro, in vivo or were certified for use along with new potential biological activities.

for the synthesis:

• Preference is given to the maximally simple reactions performed (at the level of the click-procedure) and the reactions of the poly-component condensations that allow to form easily the condensed heterocyclic systems (generally from the group of Privileged Heterocycles), containing the required substituents in the pertinent states of the heterocyclic scaffold.
• The realization of the target compounds synthesis can be carried out with the use of the modern technologies, including the synthesis in the conditions of the microwave and ultrasonic radiation, as well as the methods of the "green chemistry".

The main applied research task is to develop new efficient methods for the synthesis of medicines, colorants, and other compounds.

• The most practically feasible direction. The development of the technology and the synthesis of the substances (generic drugs) for the production of the finished pharmaceutical products. For example, there is the Order of the Russian Federation Government for the import substitution of the strategically valuable drugs. The synthesis of most of them is accessible to the staff of the laboratory. At present, the laboratory personnel conducts several projects on the development of the methodologies and the synthesis technologies of the generic drugs for a number of Russian pharmaceutical companies. Several other drugs are under discussion. The laboratory personnel has experience in the synthesis of such anticancer generic drugs as imatinib and sunitinib. The development of new high performance technologies for the synthesis of known chemical compounds and the modification of the existing technologies imply the decline in the prime cost of the products and the improving environmental performance of the processes. The synthesis of these compounds can be carried out, upon request of the customer, using the methods of the microwave and ultrasonic activation of the chemical reactions.
• More labour-intensive, but more interesting direction is the design and the synthesis of molecules of the analogues of the most effective patent medicines (circumvention of patent) as well as the search for new therapeutic properties of the drugs which are already known and authorized in clinics. Also, we are interested in the works in the field of optimization of the properties of known medicines with regard to their biotarget and the development of new molecules-analogues of known medicines which are not protected by patents.
• Own development of new non-proprietary molecules. It is a separate difficult topic for most of our pharmaceutical companies. Here the main difficulty is the lack of opportunities for conducting preclinical and clinical studies for our compounds.
• Moreover we can optimize the methodology for the synthesis of organic compounds from the class of low-molecular heterocyclic compounds in the context of developments conducted by the customer. Preference is given to blocks for Active Pharmaceutical Ingredient (API) and colorants for different purposes, including fluoresceins.

The main activities and laboratory projects in the research field of complex formation of biologically active compound with bioreceptors

Leading research worker - M. Yevstigneyev

One of the directions developed by the organic synthesis and NMR spectroscopy SRL is the study of the structure of complexes and thermodynamics of the so-called sequence specific intercalation compound of biologically active compounds with bioreceptors, in particular, with DNA.This direction was formed due to the attempt to explain the effect of changes in the biological activity of DNA-binding aromatic compounds during the injection another aromatics as caffeine, vitamin B2 (riboflavin), chlorophyllin, etc.

The main scientific problem, which scientists of NMR spectroscopy SRL try to solve, is to explain the mechanisms of biological synergism in multicomponent of DNA-binding system. The main current hypothesis, which is put forward by the laboratory personnel, is the explanation of such synergistic action of the drug mixture from the point of view of all non-covalent interactions types of aromatic compounds with each other and with DNA. The main types of such interactions are the self-association of aromatic compounds with each other (the formation of complexes of the same type of molecules), heteroassociation (the formation of complexes of the different-type molecules), and competition binding of aromatic compounds with the bioreceptor, i.e. with DNA.

This general direction of scientific activity of the NMR spectroscopy SRL is divided into four main subdirections:

• study of the energy of small aromatic molecules (ligands) complexation with nucleic acids;
• study of the structure and thermodynamics of heteroassociation of aromatic biologically active compounds (BACs);
• development of the theory of interceptor-protector action of aromatic ligands binding with nucleic acids;
• study of homo- and heterodimeric binding of biologically active compounds with nucleic acids.

The main result of the work of biomolecule NMR-spectroscopy SRL is represented by about two dozen articles published in the world's leading journals (journals with a sufficiently high impact factor), which are indexed simultaneously in two international scientometric databases, Scopus and Web of Science. The final product of the laboratory work is a new scientific knowledge about the mechanism of synergistic interaction of DNA-directed antitumor agents in combination with aromatic biologically active compounds. Theoretical developments will form the basis for fundamentally new regimens for combined chemotherapy of certain types of oncological diseases with a potentially higher suppression efficiency of both leukemia and solid tumors.

Winning of the grant of the Russian Scientific Foundation (RSF) No. 14-14-00328 "Creation of a scientific basis for a new regimen for combined fullerene chemotherapy to treat oncological diseases with DNA-directed drugs" became a stimulus in terms of achievement and overfulfillment of performance indicators for the laboratory staff. The grant was obtained in the middle of last year and was intended for the period up to 2016 inclusive. Professor Maxim Evstigneev, the Doctor of Physical and Mathematical Sciences, is the head of the project and the leading research associate of the SRL. The receipt of this grant was a logical continuation of the work begun in 2013.

As the title implies, the cornerstone of the project is the application of well-known, but currently insufficiently studied from the biological point of view, spherical carbon nanoparticles - fullerenes C60.The unique set of biomedical properties of C60 fullerene has been discovered recently, and this enables to use it as a potential element of the combined therapy of the oncological diseases. The last researches in which a laboratory personnel took part, enabled to prove exceptionally marked strengthening of antitumor effect of the doxorubicin with C60 fullerene in vitro and in vivo levels.

The works on RSF grant projects include two main parts: physico-chemical and biomedical. The physico-chemical part plans to conduct researches in interaction of C 60 fullerene aqueous colloidal solution with different bioactive compounds, it also includes detailed structured thermodynamic analysis of complex formation process, creation of the computing models and analytical approaches to study of such systems. The biomedical part studies the biological effect of fullerenes as well as the anticancer drugs in ex vivo conditions closed to native (leukemia contaminated blood), this enables to obtain the objective information about drug collaborative mechanism, to choose optimal conditions for the synergism monitoring and to formulate launch environment for conducting the first phase tests in vivo for drugs. It is worth to say that the biomedical part of the project is implemented by professors and students of the Ecology, Physiology and Biological Evolution Subdepartment of the Institute of Engineering Technology and Natural Science under the guidance of Marina Skorkina, Doctor of Biology, Associate Professor.

The main result of this project is to understand biological synergism mechanism in terms of fullerene-drug systems, and as a consequence, to create a methodology of a new more efficient process of the combined fullerene chemotherapy of some oncological diseases. This research remains the "know-how" and promises breakthrough results in improving efficiency of the modern chemotherapy. According to our scientists, the success of solving this challenge, that is launching of a new working way of combined chemotherapy in clinical experience in the nearest future, may lead to significant social and economic effect nationwide as well as worldwide and its result cannot be overestimated.

Other activities of the laboratory

Research Laboratory of NMR spectroscopy of biomolecules of Belgorod State University is equipped with modern devices for studying liquids content and the structure of dissolved substances. Instrumentation pool of the laboratory includes NMR-spectrometer 400-MR produced by Agilent (Agilent, 2012) and UVIS Ultraviolet and Visible Spectrometer Specord 210 Plus (Germany, 2014) with a Peltier attachment for regulating the temperature inside the optical cell. It should be emphasized that the NMR spectrometer is the only device of the research class in the Central Black Earth region of Russia (the nearest laboratories, equipped with similar devices are located on the periphery of Moscow-Kazan-Rostov-on-Don).

One of the activities of NMR spectroscopy SRL is the physico-chemical study of liquids, in particular, solutions of biologically active compounds. Using the method of nuclear magnetic resonance, the following problems can be solved:

• blend composition of liquids;
• studying the transformation of the liquids content in time;
• studying the physicochemical aspects of the processes in the liquid phase, including in physiological sphere.

Advantages of NMR method:

• non-invasiveness (study sample is not destroyable during measurement, in contrast to the vast majority of experimental methods);
• high power of resolution;
• high speed of analysis;
• simplicity of sampling preparation.

The NMR method can be used for studying the quality of dairy products. In this regard, two areas of applied research can be distinguished.

Branch 1 (Mixture express analysis). The object is to determine qualitative and quantitative composition of aqueous extract of milk at determining nutritive value of milk. As part of this research, the following problems could be solved:

• identification of specific components of the extract (citrate, lactate, phosphocreatine, amino acids, proteins, etc);
• mixture-ratio determination (for example lactose/protein);
• contamination test;
• disclosure of deviations, that could result from cattle condition, by comparison of given variety with reference spectrum;
• determination of qualitative and quantitative dependence of milk composition, based on diet, season, processing method, etc.

Procedure consist of spectrum measurement, further statistical analyses and drawing conclusions and recommendations. Average duration of the procedure is 30 minutes.

Branch 2 (Mixture complete analysis). Metabolical milk profiling is important developing perspective of regional, and as a whole, Russian cattle-breeding branch in terms of stock-rearing. Main purpose of the results achieved in this research is to uncover pathologies of milk donors through studying their metabolism by comparison of nuclear magnetic resonance (NMR) spectra with reference milk spectra, received by healthy donors. Besides, as part of research, followings problems could be solved:

• research of coagulative milk properties;
• calculation of number of somatic cells in milk;
• total milk quality control;
• milk components classification by different donors;
• determination of milk deterioration degree;
• determination of nutritive value of milk;
• identification of medical condition of milk donors.

It should be noted that the given branches of applied scientific researches are not high-priority for biomolecule NMR-spectroscopy SRL now. Methodology setting up period for continuous analysis is equal to 6-12 months for Branch 1 and 1-2 years for Branch 2.

Information was given by Anatoliy Buchelnikov on 30.03.2017

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