Biotechnologies

developed in Institute of Molecular Biology and Genetics of NASU

The identification of tumour-associated antigens for the development of modern immunological approaches of cancer diagnostics and treatment (Valeriy V. Filonenko, Dr.Sci., Professor).

The identification of novel tumour-associated antigens (TAA) is a prerequisite for the creation of modern anti–tumour vaccines and specific antibodies against TAA.

While using SEREX methodology (serological identification of antigens by recombinant expression cloning) more than 60 tumour-associated antigens of melanoma, colon cancer, cancer of thyroid gland and breast cancer have been identified. The listed antigens could be used in future for both diagnosis and development of novel approaches in cancer treatment.

Up-to-date a number of mouse monoclonal antibodies have been generated against TAA and signaling molecules that undergo alteration at carcinogenesis. These are antibodies to S6K1, S6K2, TSC1, TSC2, Ki67, PTEN, mTOR, CK2, FGFR1, FGFR3. These antibodies could be used for detailed diagnosis and prognosis of oncological pathologies by immunohystochemical and serological approaches.

Large-scale search for biomarkers of epithelial tumors. The creation of different diagnostic and prognostic tools for cancer research (Alla V. Rynditch, Dr.Sci., Professor, Corresponding Member of NASU).

Changes of circulating cell-free DNA levels in serum or plasma are associated with tumor burden and malignant progression. Moreover, cell-free DNA harbors genetic and epigenetic alterations specific to tumor DNA which opens the opportunity to detect tumor-specific signatures including the methylation of genes in patients serum. We evaluate the significance of cell-free DNA as a tumor marker by comparison of cell-free DNA concentration in cancer patients and healthy donors and hypermethylation determination of cancer associated genes in plasma.

We also investigate the expression and function of possible predictive marker for breast cancer – intersectin 2 (ITSN2). Intersectin 2 has been proposed to be used in diagnostic tests for prediction of disease outcome after breast cancer surgery and CMF-based chemotherapy. We analyze expression profiles of ITSN2 alternatively spliced isoforms in different groups of breast cancer patients and investigate the complex protein-interaction network of adaptor protein ITSN2. The results obtained during this project are expected to be helpful for the understanding of tumor progression mechanisms and will make a contribution to the approval of intersectin 2 as one of breast cancer predictive markers which could find clinical application.

From high-throughput analysis to creation of marker panel for diagnostics of different epithelial tumors (Volodymyr I. Kashuba, Dr.Sci., Senior Research Scientist).

In Department of Molecular Oncogenetics the microarrays module was created that consists of tools for the designing and application of DNA microarrays. In collaborative work with Karolinska Institutet (Stockholm, Sweden) was developed NotI-microarray technology that allows performing global searching for genetic and epigenetic alterations of genes from chromosome 3 in epithelial tumors. The main advantages are simultaneous detection of genetic (deletions or amplifications) and epigenetic (methylation and demethylation) alterations. In current work we expanded an application of NotI-microarray from single chromosome to whole genome scanning of hot regions – genes which demonstrate altered expression in epithelial tumors (kidney, prostate, colon etc.). We use the dualscan procedure which includes cross-platform analysis of gene expression with subsequent location of NotI-sites through human genome. It allows us to isolate genes which are used to design NotI-microchip for profiling hot regions in epithelial tumors. Subsequent application of selected set of genes is the creation of panel for detection of epithelial tumors by means identifying nucleic markers in body fluids.

The development of complex molecular diagnostics of chronic myelogenous and acute lymphoblastic leukemias using PCR and specific polyclonal antibodies (Stanislav S. Maliuta, Dr.Sci., Professor, Corresponding Member of NASU; Gennadiy D. Telegeev, Dr.Sci., Senior Research Scientist).

The method was tested on more than 200 patients from different regions of Ukraine after the corresponding assignment from The Institute of Hematology and Transfusiology of the Ministry of Public Health of Ukraine and from regional hematological departments. Entire correlation with other diagnostic tests and high sensivity (10–4–10–5 of malignant cells) were obtained. This method gives the possibility to detect the type of translocation as well as to exclude false results completely. It is very important to estimate prognosis and therapy protocols.

Identification of transcryptome changes and detection of signatures for human brain tumor classification and diagnostics (Vadym M. Kavsan, Dr.Sci., Professor, Corresponding Member of NASU).

Identification of changes in gene expression, potentially valuable for diagnostics and prognostics of glial tumors based 89 on hybridization analysis of DNA microarrays and serial analysis of gene expression (SAGE) using various statistical methods including artificial neural network. The selected genes, valuable for prognosis, will allow identifying gene expression signatures, suitable for molecular typing of glial tumors and prognostic evaluation of this devastating disease.

Nanoconjugates of natural biopolymers with antisense oligonucleotides for inhibition of glial tumors (Vadym M. Kavsan, Dr.Sci., Professor, Corresponding Member of NASU).

The developing of new polyfunctional nanoconjugates for specific targeting the increased production of specific proteins associated with proliferative and angiogenic properties of tumor cells. Specific short interfering RNA (siRNA) or antisense morpholino oligonucleotides to CHI3L1 and VEGF mRNAs, as well as bradykinin antagonists or azolidinone derivatives and known anticancer chemotherapeutic drug temozolomide in different combinations will be attached to polymer matrix, polymaleic acid (PMLA). Drug-releasing unit contains the antibody against transferrin receptor (TfR) to specifically target tumor cells and disulfide bonds, which are stable in the bloodstream and endosomes/lysosomes but are naturally cleaved by cytoplasmic glutathione.

Plasma cell-free DNA as diagnostic and prognostic tools for renal, colon cancers and metastatic uvenal melanoma (Inessa Ya. Skrypkina, Ph. D., Senior Research Scientist).

The possible use of plasma cell-free DNA as an important source for detection of methylation status of sets of tumorsuppressor genes, tumor-specific mutations and levels of cell-free DNA concentration for determination early stages of oncological diseases are under investigation. Methodolodies of efficient isolation of cell-free DNA from plasma, and evaluation of its concentration by quantitative PCR assay, determination of methylation status of CpG islands of selected genes associated with development of renal and colon cancers as well as detection of mutations specific for metastatic uvenal melanoma by PCR combined with sequencing of the fragments specific for several genes associated with this disease are used for creation of cell-free DNA-based test systems. The advantages of such systems are: high sensitivity and specificity due to the usage of large number of tumor marker genes, the possibility to use the system for early diagnostics of cancer, noninvasive sampling methodology, no need in expensive equipment.

The establishment and operation of a laboratory module for microarray fabrication and application (Volodymyr I. Kashuba, Dr.Sci., Senior Research Scientist).

  • Diagnostics of different pathologies, especially, malignant tumors
  • Early diagnostics of tumors of different localization sites.
  • Prognosis of the course of cancer disease and the response on therapy
  • Diagnostic of pathogenic microorganisms

Test systems for gene diagnostics of severe hereditary diseases and hereditary susceptibility to mass pathologies (Ludmila A. Livshits, Dr.Sci., Professor).

The elaboration of diagnostic methods and individual components of the kits for DNA analysis of mutant genes that cause the development of the most widespread monogenic hereditary diseases in Ukraine (cystic fibrosis, phenylketonuria, spinal muscular atrophy, Duchenne muscular dystrophy, Martin-Bell syndrome (fragile X), haemophilia A, Huntington’s disease, hereditary hemochromatosis, Charcot-Marie-Tooth neuropathy (CMT), hereditary corneal dystrophies, Stargardt’s macular dystrophy), hereditary susceptibility to mass pathologies (cardiovascular, reproductive, inflammatory, endocrine) and genetic forms of men (azoospermia, oligospermia) and women (premature ovarian failure) infertility is being held. These diagnostic kits can be used for genetic testing, that is held during prenatal diagnostic, presymptomatic diagnostic, specification of the diagnosis, selective and mass screening for heterozygous carriers in the high risk groups and in the general population. The results of the genetic testing are the necessary requirements for prevention and precise treatment for the improvement of the demographic setting in Ukraine.

The list of the institutions, where the elaborated methods of the DNA-analysis for the clinical diagnostics of patients with cystic fibrosis, phenylketonuria, Martin-Bell syndrome (fragile X), hereditary hemochromatosis and secondary pathologies connected with it, Huntington’s disease, Charcot-Marie-Tooth neuropathy, hereditary corneal dystrophies, genetical forms of men and women infertility, were used in: State Institution “Institute of Pediatrics, Obstetrics and Gynecology of NAMS of Ukraine” (Kyiv); State Institution “National Research Center for Radiation Medicine of NAMS of Ukraine” (Kyiv); State Institution “Institute of Neurosurgery named after A. P. Romodanov of NAMS of Ukraine” (Kyiv); State Institution “V. P. Filatov Institute of Eye Diseases and Tissue Therapy of NAMS of Ukraine” (Odessa); State Institution “Institute of Occupational Medicine of NAMS of Ukraine”(Kyiv); Institute of Hereditary Pathology, NAMSU (Lviv); Crimean Republican Specialized Medical- Genetic Center (Symferopol); Ukrainian Center of Diagnostics and Treatment of Cystic Fibrosis (Odesa); Interregional medical genetic centers in: Kyiv, Donetsk, Kharkiv, Odesa, Kryvyi Rig; Regional medical genetic consultations: Ternopilska, Rivnenska, Poltavska, Khmelnytska, Vinnytska, Volynska, Ivano-Frankivska, Chernigivska, Zakarpatska, Luganska; Clinic “Isida-IVF” (Kyiv).

Test system prototypes have been developed for the DNA diagnostics of following hereditary monogenic diseases: cystic fibrosis, phenylketonuria, spinal muscular atrophy, Martin-Bell syndrome and hemochromatosis.

We obtained the patent for utility model N64654 Method for DNA diagnostics of DF508 mutation in cystic fibrosis transmembrane regulator protein gene (CFTR) / O. O. Soloviov, S. A. Kravchenko, L. A. Livshits, owner IMBG NASU - ¹ u 2011 05507, publ. 10.11.2011, Bulletin ¹ 21.

The structural–based drug discovery against pathogenic bacteria (Mykhaylo A. Tukalo, Dr.Sci., Professor, Corresponding Member of NASU).

We are using the differences between human and prokaryotic enzymes for the development of selective inhibitors as potential drugs against Mycobacterium tuberculosis, Enterococcus faecalis and Streptococcus pneumonia. The search strategy for antibacterial compounds is based on the combination of X-ray structural analysis of the target protein, computer modeling of the interaction of low-molecular ligands with the target protein and synthetic procedures of combinatorial chemistry.

Using flexible docking technique, the virtual screening of 100000 compounds from in-house collection (at the Department of Medicinal Chemistry) was carried out, and after in vitro and in vivo biological screening several inhibitors of M.tuberculosis leucyl-tRNA synthetase with anti-TB activity were selected.

The production of a novel antiangiogenic cytokine cendomap – C-module of tyrosyl-tRNA synthetase (Olexander I. Kornelyuk, Dr.Sci., Professor, Corresponding Member of NASU).

A novel biotechnological product – cendomap cytokine has been obtained and tested in vitro. The bacterial system for the expression and purification of cendomap cytokine permits to acquire a recombinant protein at preparative scale of about 10 mg from 1 liter of bacterial culture. Cendomap cytokine may be widely used as a mediator of procoagulant and antiangiogenic action, an inductor of apoptosis and, in perspective, as a putative antitumour medication for clinical oncology.

Development of a new multiplex PCR-based test for quick screening of staphylococci and methicillin-resistance in hospital patients with sepsis (Olena V. Moshynets, Ph.D., Research Scientist).

Sepsis and its after effects are currently significant problems in hospital medicine. Sepsis is a potentially deadly medical condition characterized by a whole-body inflammatory state (so-called systemic inflammatory response syndrome) caused by severe infection. The early identification of sepsis and its effective treatment are especially important in some groups of patients, including new-borns, cancer and intensive care patients. Successful treatment depends on the early detection of the etiological agent of the sepsis as well as of any antibiotic resistances. Bacteria belonging to Staphylococcus spp. are the most important Gram-positive causative agents of sepsis and are responsible for ~ 50 % of sepsis and pyoinflammatory diseases in Ukraine. Up to 70 % of Ukrainian staphylococci are also methicillin resistant which is determined by the mec-containing cassette SCCmec encoding the mecA β-lactam resistance gene. For this reason, a rapid screening assay is vitally needed in Ukraine for both the early identification of staphylococci and methicillin-resistance. Two PCR-based assays for Staphylococcus spp. and four for methicillin- resistance identification were tested using forty Staphylococcus spp. isolates obtained from patients of the Ukrainian Research Institute of Traumatology and Orthopedics, Ministry of Health of Ukraine (Kyiv). Based on the results of this work, a new “in house” multiplex PCR test for the direct simultaneous identification of Staphylococcus spp. and methicillin resistance was developed.

The molecular modeling and drug design: development of protein kinase inhibitors (Sergiy M. Yarmoluk, Dr.Sci., Professor).

Nowadays, drug development starts from identification of biological macromolecule target (DNA, or, in most cases, enzyme) responsible for certain disorder. By altering target activity with drug-like organic compounds it is possible to prevent or treat target-associated disorder(s). Among human proteins, protein kinase family has one of the greatest medicobiologic meanings.

Protein kinases constitute at least 2 % of the human genome and represent one of the most fundamental intracellular signaling mechanisms. Imbalances in kinase activity are a major factor in many disease states, especially those involving inflammatory and proliferative responses. Hence, protein kinases are attractive targets for drug research.

The goal of proposed project is the development of protein kinase inhibitors.

The approach used to develop protein kinase inhibitors is based on combined application of computer modeling techniques, directed organic synthesis and biological in vitro screening. The main points of the project are:

  • Virtual screening of combinatorial libraries for hit compounds using docking and molecular dynamics simulation techniques, homology modeling (if required)
  • Chemical synthesis of promising compounds
  • In vitro testing of synthesized compounds and QSAR
  • Lead compound identification
  • Lead optimization

We have been developing the inhibitors of Casein Kinase 2 (CK 2) for about a year. At the time we have founded 3 lead compounds with IC50 up to 10–7 M and now perform their chemical optimization.

Telomerase inhibitors based on specific ligands of quadruplex DNA (Igor Ya. Dubey, Dr.Sci., Senior Research Scientist).

Telomeric DNA and telomerase are currently considered promising targets for cancer therapy. Telomeres are guaninerich DNA sequences located at the ends of the chromosomes that can adopt a specific quadruplex structures based on the stacks of G-quartets. Tumor cells express high levels of telomerase enzyme responsible of maintaining telomere length, whereas normal cells are devoid of telomerase activity. Small molecules that specifically bind to quadruplex structures 91 of telomeric DNA can inhibit telomerase in cancer cells and thus demonstrate antitumor properties.

We have prepared a series of novel cationic porphyrin derivatives, their metal complexes and conjugates as efficient G-quadruplex binders. Some of these compounds have been shown to inhibit telomerase activity in vitro at micro- and even nanomolar concentrations (TRAP assay). Metal complexes of cationic porphyrins conjugates with intercalating agents demonstrated antiproliferative activity in tumor cell culture with IC50 values in the range 6-11 μM comparable to those of doxorubicin and vincristine.

Antimycobacterial agents (Anatoly D. Shved, Dr.Sci., Senior Research Scientist).

Sharp increase in human morbidity and mortality caused by tuberculosis becomes a worldwide problem. It is mainly associated with Mycobacterium tuberculosis strains which are resistant to one or more common therapeutic agents. We have synthesized the library of new aryl and hetaryl amides of phenazine-1-carboxylic acid as potential antimycobacterial agents capable to inhibit RNA synthesis in vitro. Their biological activity has been investigated in collaboration with Italian research team led by Prof. A. De Logu. A range of synthesized compounds have inhibited the growth of M.tuberculosis strains in vitro at concentrations equal or even lower than those for well-known antituberculosis drugs isoniazid, rifampicin and ethambutol. Moreover, some derivatives have shown high activity against clinical isolates with multiple drug resistance.

The baculovirus expression system for obtaining recombinant proteins (Olexander P. Solomko, Dr.Sci., Professor).

The system for an expression of recombinant proteins on the basis of a virus of nuclear polyhedrosis Malacosoma neustria (Mane) and cultures of insect cells was designed in the department of biochemical genetics. The designed system is the first baculovirus system of an expression of recombinant proteins in Ukraine. At present the similar systems find wide application in the world for obtaining recombinant proteins, including for effective antiviral vaccines. The laboratory procedure of obtaining biologically active recombinant human Prolactin with our system and commercial system was designed in cooperation with the Institute of biological chemistry and biophysics (Poland). The method is covered by the patent of Poland. Human Prolactin is a multifunctional hormon, which participates in many physiological processes. Recombinant Prolactin is necessary for effective tests – systems for detection of some pathologies (ischemia, renal failure, brain tumour). The protein can be used in therapy.

New anticancer and antiviral medications (Anatoliy I. Potopalsky, Ph.D., Docent).

Medications amitozyn and izatizon with antiviral, antitumor and immunoregulating activity have been elaborated.

Amitozyn manifests antitumoral action and antimetastasis activity, but in comparison with existing medications it does not depress the hematosis processes, also it does not cause allergic reactions and has low toxicity. Amitozyn has undergone clinical tests in the system of the Ministry of Health of Ukraine. Izatizon has been approved in veterinary institutions and is studied in medicine. By using elaborated medications the technologies of diagnostics, prevention and treatment of plants agrobacterial cancer, non-vaccine prevention and treatment of mass virus diseases, increase in productivity of plants, mushrooms, useful insects (such as bees, silkworms, and oak silkworms), fish, poultry and animals have been worked out.

Based on the new biotechnology of the accelerated obtaining of new plant forms and varieties using nucleic acids preparations, such sorts as the feed lupine “Industrialny”, pumpkin “Kavbuz Zdorovyaga”, purple echinacea “Poliska Krasunya”, rye “Drevlyanske”, potato “Dzvin”, tomato “Ukrainski” were obtained and registered in the State Register for the Plant Varieties of Ukraine. The elaborations give the significant social and economic effect.

Biotechnology of the plant secondary metabolites production (Stanislav S. Maliuta, Dr.Sci., Professor, Corresponding Member of NASU).

An efficient method for the increase of Glycyrrhizin (Gl) and flavonoids production from Ri-plasmids transformed licorice Glycyrrhiza sp. cells and hairy roots was developed. An endophyte mycorrhizal fungus was isolated from intact roots of the medical plant Potentila alba L. as a potential elicitor to stimulate the total biosynthesis of the secondary metabolites in transformed licorice cells and roots by 1.5 fold. Glycyrrhizin (triterpen saponin) production in the biotechnological way is very perspective as a source of the natural sweetener for the diabetes patients, and its aglycone Glycyrrhizic acid has shown the inhibiting II-β-dehydrohynase (hepaprotective effect in liver and kindneys tissues in vitro as well as in vivo). Flavonoids, 18-dehydroglycyrrhizic acid and Glycyrrhizinic acid have shown antioxidation effect. It is their anti-inflammatory effect by which one can explain their positive therapeutic effect on lungs and liver irradiated impact pathology, and note the anti-sclerotic, anti-inflammatory, anticancer, and anti-ulcer activity of licorice metabolites.

The application of lectins from medicinal plants for biomedical research (Iryna S. Karpova, Dr.Sci., Senior Research Scientist).

The lectins are a large group of carbohydrate-binding proteins, which can specifically recognize receptors exposed on cellular membranes in all types of living organisms. In humans endogenic lectins play their role in fertilization, immune response, antiviral and antibacterial protection, clearance, apoptosis etc. Plant exogenic lectins have been widely used as a tool in the investigation of animal membranes which differ in their surface sugars: in case of immuno-haematological and endocrine diseases, developing infection or neoplastic process and other pathological changes. In the United States some lectins are commercially used for purging of bone marrow for transplantations in leukemia patients. An original technology of medicinal plant lectins isolation and purification has been developed. The new approach to receptor diagnostics of different pathologies (such as acute radiation sickness and thyroid gland diseases) using a panel of lectins isolated from medicinal plants in reaction of haemagglutination with data processing has been proposed. The method is based on the unique properties of lectins to indicate the deviations in membrane structure. The endogenous misbalance of lectins occurs in a pathological state and the lectino-test as its indicator has been proposed. The application of herbal lectins in diagnostics and individual approaches to therapy is being developed. Contemporary research is devoted to further studying of peculiarities of medicinal plants lectins: their adhesive, immunomodulating and antitumour activity, and possible usage for stem cell identification and separation.

Unique cell strains of rare medicinal plants have been generated that produce bioactive compounds important for medicine and pharmacology (Victor A. Kunakh, Dr.Sci., Professor, Corresponding Member of NASU; Oksana O. Poronnik, Ph.D.; Ludmila P. Mozhylevska; Volodimyr I. Adonin).

Technology for production of Rauwolfia serpentina cultured cells with high content of ajmaline, that is used as a basic compound for antiarrhythmic and antihypertensive drugs. The technology for production of biomass with ajmaline content of about 1.0 % was tested under the industrial conditions.

Technology for biomass production and micropropagation of Ungernia victoris, perennial plant, endemic of Pamir that is used as a source of alkaloids galantamine and licoryne. Preparations based on galantamine are used for treatment of myasthenia progressive muscle dystrophy (myopathia), radiculitis. Licoryne derivatives are used to treat the chronic and acute inflammations of respiratory system, and bronchial asthma.

Technology for Panax ginseng cell biomass production for use in the manufacturing of medicines, nutritional supplements and cosmetics. Techniques have been developed for regulation of ginsenosides biosynthesis as well as their quantitative and qualitative composition in callus biomass.

Technology for production of Echium plantagineum cultured cells biomass with high content of shikonin, which is used as a color additive, medical remedy with antitumor, antibacterial, anti-inflammation, wound healing and antiviral activities, and a compound of cosmetics with medicinal properties.

A new test system “Escherichia coli – bacteriophage λ” have been developed to study the mutagenic and antimutagenic properties of biologically active compounds (V. A. Kunakh, T. P. Pererva, H. Yu. Miriuta, L. P. Mozhylevska).

Nanocellulose-based composites (Anatoliy I. Potopalsky, Ph.D., Docent; Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist)

Antimicrobial biodegradable composites based on nanocellulose and antimicrobials (isatison; polyhexamethylenguanidine chloride, PHMG-Cl) for use in medical practice have been designed. New composite of nanocellulose with multipurpose drug izatizon might be effective for a treatment of wound surfaces, ulcers and burns, melanoma, as well as against herpes and other viral infections of the skin. The PHMG-Cl prevents the biofilm formation by microorganisms and could be further exploited as a wound dressing for healing and the regeneration of a scarless skin.

A concept of low-cost technology of growing and utilizing pioneer plants in a lunar garden (Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist).

On the base of the model system Tagetes patula L. – anorthosite (analog of lunar soil) – a consortium of microorganisms, a concept of low-cost technology of growing and utilizing pioneer plants in a lunar garden has been developed for the first time. The putative scenario anticipates using a local material – regolith – and a consortium of rationally assembled microorganisms (biomobilizing plant-essential elements, plant growth promoting, plant protection from abiotic stressors, etc.).

Test-system for deteñtion of potato viruses VPX, VPY, VPM, and viroid by RT-PCR (Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist).

Specific for conditions of Ukraine phytoviruses VPX, VPY, VPM and viroid have been isolated on the basis of potato disease symptoms and proved by ELISA and ÒEM. A set of the test system consists of primers pairs specific to the gene, encoding virus capsid protein, designed for either phytovirus, and positive controls, fragments of capsid protein, gene have been cloned in plasmid. The set for phytoviruses detection was tested at the Institute for Potato Research of the National Academy of Agricultural Sciences, NAASU (t. Nemishaeve, Kyiv region).

Bacterial inoculant KLEPS® (Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist).

The first bacterial inoculant KLEPS®, which provides the plant with biological nitrogen, growth promoters and enhances its defense system, has been designed and registered in Ukraine. The inoculant is recommended for pre-treatment of seeds of cereal and vegetable crops. KLEPS® increases yield by 10–40 %, forms an extensive root system that provides early flowering and ripening crops, reduces the incidence of cultures. Application of KLEPS® reduces the use of mineral nitrogen fertilizers and pesticides by 70 % and allows protection of the environment.

Express method of the Klebsiella oxytoca authentication by PCR (Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist).

Unique sequence of the pehX gene we cloned from industrial strain of bacterium K. oxytoca IMBG26 enabled us to develop a method of the bacterium identification by polymerase chain reaction. Rapid, sensitive and specific test is recommended for discrimination of K. oxytoca between similar types of bacteria of the genus Klebsiella, as well as for the environmental monitoring of K. oxytoca.

Technology of the use of bacterial inoculants for adaptation of potato in vitro to post vitro conditions and protection of plants from pathogens in the commercial production of potatoes (Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist).

The principles of the use of biological products that contain beneficial bacteria for plant protection from pathogens in the process of adaptation of potato in vitro plants to post vitro conditions have been elaborated.

Technology of bacteria-assisted phytostabilization of heavy metals (Nataliia O. Kozyrovska, Ph.D., Senior Research Scientist).

To implement the plant-microbe system to a variety of polluted environments for a remediation, a flexible and hardy plant-bacteria system is designed. A rationally assembled bacterial consortium for a multipurpose application at different level of heavy metal (HM) pollution in the soil is proven in the field model and natural environments. In the case of aerogenic HM pollution (transport and industrial off-gases) bacteria promote growing clean crops. On the other hand, the consortium provides environmentally appropriate technology for HM phytostabilization on moderate and extremely polluted lands near industrial enterprises and mines.

Gene and Cell Therapy (Vitalii A. Kordium, Dr.Sci., Professor, Corresponding Member of NASU, Full Member of NAMSU).

The main directions of the department activity are the study of fundamental basis of aged and sick organism renewal, the development and usage of gene and cell technologies for damaged cells and hereditary material replacement, as well as biotechnological obtaining o therapeutic recombinant proteins.

Approaches to the obtaining of mesenchymal stem cells from Wharton jelly for further clinical usage were developed. MSC were characterized morphologically, by surface and inner markers, ability to differentiate in first 6 passages. It has been shown that practically all the characteristics change after the 3rd passage in cell culture confirming that only 1-2 passages answer the requirements for the clinical use.

Scientific approaches for gene therapy technologies of multifactor diseases have been developed. They are based on the introduction of recombinant genetic material into an organism with the aim of pathologies correction. Effective non viral systems for target therapeutic genes delivery to the cells and organs were created. The systems for the targeted therapeutic genes delivery into specific cells and tissues that will ensure high-level transgene expression in organism are developed. Positive results of gene therapy for pathologies, namely artificially induced insulin – dependent diabetes mellitus and atherosclerosis – were obtained on laboratory animals.

Today, the most promising way to deliver in organism different cells, which are the source of therapeutic agents (hormones, growth factors, cytokines), is the encapsulation of such cells in semipermeable microcapsules or another devices. In the scope of department research the approaches to the delivery of genetically engineered cells that produce several cytokines (LIF, FGF-2, IL-10) are developed. Significant success was achieved in genetic engineering biotechnology. The scientists of the department developed the unique method of recombinant proteins obtaining in E. coli with the use of λ phage. This method has no world analogues, it provides for target product obtaining in high concentration in active soluble form directly in culture medium. Its effectiveness for obtaining proteins of both pro- and eukaryotic origin was shown. Industrial technologies for such products as ß-galactosidase, human α-2a and α-2b interferon, human growth hormone, etc. obtaining were developed according to this method. Preparation of recombinant human α-2a interferon, under the trademark “Laferon” is being produced in Ukraine for more than 10 years, and is successfully used in medical practice.

Today the problem of ischemia of renal parenchyma is an important issue for diagnostics and development of further therapeutic treatment of patient with renal pathology. The experimental data obtained revealed that the injection of complex of bFGF with our developed carrier (that increases its half-life in tissues) led to the enhancement of blood perfusion in ischemic kidney under condition of chronic ischemia and protects contralateral kidney from the development of irreversible sclerotic changes.

Theoretical basis of damaged hereditary material substitution due to exchange of genetic material between cells, including stem cells, is developed. Now it is under experimental examination.

Development of technology for repairing injured liver function by MSC transplantation (Svetlana Yu. Rymar, Ph.D., Senior Research Scientist).

Liver disease is one of the leading death causes (4th rank) in Ukraine. In the world Ukraine occupies 9th rank in the liver disease mortality. Currently orthotopic liver transplantation represents the only therapeutic option for patients with advanced liver diseases and hepatic failure. Now the cell therapy has been proposed as an alternative way to treat liver injury. Chronic liver injury induces liver fibrosis resulted in cirrhosis terminated with hepatocellular carcinoma. In this investigation the process of liver injury causes by CCl4 to 94 laboratory rats. The characteristics of cirrhosis development in rats are similar to human cirrhosis. Human cord MSC isolated and cultivated in our department may be used for the transplantation. Based on the liver injury dynamics data, a detailed protocol of human cord MSC transplantation will be developed. The protocol will result in an effective rescue experimental liver failure and contributes to a liver regeneration and offers a potentially alternative therapy for treatment of liver diseases.

The development of skin equivalents on the base of human stem cells cultivated on special carriers in vitro for treatment of massive burns (Lyubov L. Lukash, Dr.Sci., Professor).

The biotechnology of skin equivalents obtaining with the usage of adult human mesenchymal stem cells has been developed (patent of Ukraine UA N 82583). Particularly the database of characterized cell lines originated from samples of embryonic and adult human tissues and compositions of cultural mediums, carriers and hydrogel substrates applied at the production and optimization of skin transplantants is in progress. There is also in progress of creating a cellular bank, which will be connected with Euro Skin Bank as a central provider of biomaterials for treatment of burns in the USA and Europe, and the international organization Bone Marrow Donor Worldwide which supplies the transplantations of bone marrow. The joint activity with the Center of Thermal Burns and Plastic Surgery of the Ukrainian Ministry of Health has been initiated on the introduction of the developed by the Institute skin equivalents, based on human fibroblasts, into medical practice.

The development and creation of electrochemical and optical biosensors for medicine (Anna V. El’skaya, Dr.Sci., Professor, Full Member of NASU; Alexei P. Soldatkin, Dr.Sci., Professor, Corresponding Member of NASU).

  • Development of potentiometric mono- and multibiosensors specific to glucose, urea and creatinine for the medical diagnostics and monitoring of hemodialysis
  • Development of amperometric microbiosensors for in vivo determination of main metabolites (glucose, lactate,ATP) and monitoring of some neurotransmitters (acetyl choline, choline, glutamate and D-serine) in brain of mammals
  • Development of immuno- and DNA-sensors based on surface plasmon resonance for revealing some mutations and pathogenic microorganisms.

The development and creation of analytical devices based on electrochemical mono- and multibiosensors for environment monitoring (Anna V. El’skaya, Dr.Sci., Professor, Full Member of NASU; Tatiana A. Sergeeva, Dr.Sci.; Yaroslav I. Korpan, Ph.D.).

  • Development of potentio- and conductometric enzyme mono- and multibiosensors for direct and inhibitor analyses of single toxicants and their mixtures (herbicides, pesticides, steroidal glycoalcaloids, formaldehyde, heavy metal ions, etc.)
  • Development of technology of production of synthetic analogues of biological receptors (polymers-biomimics) and creation on their basis of a number of electrochemical sensors (conductometric, capacitive and amperometric) for determination of herbicides of triazine type.

The development and creation of electrochemical biosensors for biotechnological processes and foodstuff control (Alexei P. Soldatkin, Dr.Sci., Professor, Corresponding Member of NASU; Sergiy Dzyadevych, Dr.Sci., Professor).

  • Development of amperometric biosensors and sensors arrays for determination of glucose, glycerol, ethyl and lactate in wine samples
  • Development of potentio- and conductometric biosensors for penicillin determination
  • Development of the potentiometric enzyme sensor based on inhibitor analysis for determination of nature neurotoxic substances – glycoalcaloids – in potato, tomatoes and other foodstuff of solanaceous type
  • Development of conductometric biosensors and sensors arrays for determination of carbohydrates in food industry.