Laboratory of Biosynthesis of Nucleic Acids

The Laboratory was established on the basis of the Department of biosynthesis of nucleic acids (former head Corresponding Member of NASU, Prof. Kavsan V.M.).

Head

Inessa Ya. Skrypkina

Ph.D. (Mol. Biol.), Senior Staff Scientist
: (380-44) 526-34-98;
: (380-44) 526-07-59;
E-mail: i.skrypkina@imbg.org.ua

Education and Degrees:

19841989 Graduate Student, Faculty of Biology, Taras Shevchenko National University of Kyiv, Ukraine, M. Sc. (molecular biology), Kyiv, Ukraine

2003 Ph. D. (molecular biology). Thesis: "Characterization of novel genes on human chromosome 21 and identification of their homologues in mouse"

2006 Senior Researcher (molecular biology)

Professional Employment:

19892001 Engineer, Department of Biosynthesis of Nucleic Acids, Institute of Molecular Biology and Genetics (IMBG), NASU, Kyiv, Ukraine

2001-2004 Junior Research Scientist, IMBG NASU, Kyiv, Ukraine

20042006 Research Scientist, IMBG NASU, Kyiv, Ukraine

20062017 Senior Research Scientist, IMBG NASU, Kyiv, Ukraine

2017 Leading Research Scientist, IMBG NASU, Kyiv, Ukraine

from 2018 Head of the Laboratory of Biosynthesis of Nucleic Acids, IMBG NASU, Kyiv, Ukraine

Membership:

since 2007 Member of FEBS (Federation of European Biochemical Societies)

since 2003 Member of Vavilov Society of Geneticists and Breeders of Ukraine (VSGBU)

Research Area:

Determination of genetic and epigenetic changes in extracellular nucleic acids of biological fluids of patients with kidney cancer as the diagnostic and prognostic markers of early noninvasive diagnostics

Investigation of the mechanisms of malignant tumors initiation and progression in order to create the general therapeutic approaches to cancer treatment

Clarification of the role of potential oncoproteins interactions and tumor suppressor proteins in RAS/MAPK and PI3K/AKT signaling cascades, involvement of these interactions in the malignant transformation of brain cells, proliferative and invasive properties of tumor cells and search for specific inhibitors of these signaling pathways

Determination of the role of the endogenous retrovirus gene env (ERVW-1) in the interaction of cancer cells with microenvironment and in the regulation of carcinogenesis, which will allow estimating the predictive value of ERVW-1 in the treatment of cerebral and colon cancer. The experimental data obtained will serve as a fundamental basis for the development of new promising therapeutic approaches to cancer treatment.

urrent Research Activities:

Hundreds of genes expression changes have been revealed in tumors of glial and meningial origin by modern methods of expression genetics. Opposite changes of several genes expression suppose different mechanisms of these tumors development and can be used as molecular biomarkers. 129 genes with 5-fold changed expression were found in glioblastoma, the most aggressive human brain tumor. 44 of them were overexpressed genes, which participate in angiogenesis, immunity, ECM, cell signaling pathways, and related to the IGF-system. IGF1 is a key peptide in many tumors but its gene was not found as overexpressed in glioblastoma. It was shown that CHI3L1 gene with considerably increased expression could participate instead of IGF1 in the development of glial tumors.

The new human cell line stably producing CHI3L1 was constructed and found that these cells had an accelerated growth rate and could undergo anchorage-independent growth in soft agar that is one of the most consistent indicators of oncogenic transformation (Fig. 1).

Fig. 1. Cells with overexpressed CHI3L1 oncogene formed significantly more colonies in soft agar which is a sign of malignant transformation. A 293 cells stable transfected by a pcDNA3.1_CHI3L1 plasmid which expresses CHI3L1 gene. B 293 cells stable transfected by empty pcDNA3.1 plasmid vector.

293_CHI3L1 cells had activated PI3K and MAPK pathways; phosphorylated AKT was localized in cytoplasm, while ERK1/2 were localized in both cytoplasm and nuclei where they could activate different transcription factors with certain biological outcome. The formation of tumors in rats by 293 cells expressing CHI3L1, evidenced that CHI3L1 is an oncogene which is involved in tumorigenesis. It was the first animal model of human brain tumor which could be used for studying of various biological properties of brain tumors in the immunocompetent animals (Fig. 2).

Fig. 2. Malignant tumors in the rat brain after stereotactic intracerebral implantation of 293_CHI3L1 transformed cells. A 293_CHI3L1 cells were implanted under ketamine anesthesia in kaudoptamen using Narishige stereotactic device, according to the coordinates of Swansons Brain Atlas. B General view of tumor paraffin section, initiated by 293_CHI3L1 cells. Hematoxylin-eosin staining

It was found that CHI3L1 gene promotes chromosomal instability. Constitutive expression of CHI3L1 leads to qualitative and quantitative chromosomal abnormalities, contributes to the malignant phenotype by accelerating cell proliferation, and also increasing the genetic heterogeneity of cell populations (Fig. 3).

Fig. 3. Constitutive expression of the oncogene CHI3L1 promotes chromosomal instability in cells. Karyographs of the cell lines 293 and 293_CHI3L1 clone 2. X axis designates the chromosomes, axis Y chromosomes copy number, axis Z quantity of karyotyped cells (20 cells). Karyographs demonstrate variability and clonality of chromosomal changes within cell lines

CHI3L1 gene knockdown by CHI3L1 siRNA transfection gave noticeable CHI3L1 protein blockade (80-90 %) with significantly reduced. pERK1/2 and the colony-forming ability of 293_CHI3L1 cells in soft agar (Fig. 4). The obtained results demonstrate that activity of CHI3L1 mediated by pathways involved ERK1/2 and AKT has a growth-promoting role during tumorigenesis and indicate that efforts to inhibit its activity should be considered during cancer therapy.

Fig.4. Western blot analysis displayed CHI3L1 gene knockdown in 293_CHI3L1 cells

Along with superexpressed genes, it was found 85 genes relating to the potential tumor suppressor genes. The results show that CHI3L2 is an antagonist to CHI3L1 and if CHI3L1 is a real oncogene that may play an important role in tumorigenesis, CHI3L2 is anti-oncogene. A spatial model of CHI3L2 protein was constructed and it was revealed the main structural features that distinguish it from the homologous one but functionally opposite CHI3L1 protein (Fig. 5). Heparinbinding site of CHI3L1 has been identified using site-directed mutagenesis and it has been shown that it might be responsible for the oncogenic properties of CHI3L1.

Fig. 5. Three-dimensional structure of CHI3L2 protein obtained by a modeling on the base of homology with CHI3L1 protein. A CHI3L1 protein. B CHI3L2 protein

The antiproliferative properties of two distinct classes of molecules, namely bradykinin (BK) antagonists and azolidinones, were shown in three different in vitro models of malignant transformation: 293 cells, stably transfected by CHI3L1 oncogene (293_CHI3L1), human glioblastoma cells U373 and mantle cell lymphoma (MCL) cells Granta, JeKo, Mino, and UPN1 (Fig. 6). For drugs delivery into the brain tumors are used the nanocojugates of Polycefin on the basis of polymaleic acid which can penetrate across the blood-brain barrier.

Fig. 6. Inhibition of 293_CHI3L1 and U373 cells growth by bradykinin antagonists

Artificial intellect approach was used for diagnostics of glial brain tumors by self-organized Kohonens map (SOM). Obtained data clearly show the clusterization of glioblastoma and normal brain samples (Fig. 7).

Fig. 7. Classification of brain tumors using artificial neural network. Distribution of glioblastoma and normal brain samples using Kohonen map

National Grants:

Projects of National Academy of Sciences of Ukraine:

  • 2015-2019 Complex targeted multidisciplinary program of the scientific research of the NAS of Ukraine Fundamental basis of molecular and cellular biotechnologies. Project Identification of genetic and epigenetic changes in the extracellular blood DNA of patients with kidney or breast cancer, as the diagnostic and prognostic markers of early non-invasive diagnostics (0115U002951) of National Academy of Sciences of Ukraine (scientific supervisor Skrypkina I.).
  • 2017-2018 Young scientists research projects of the National Academy of Sciences of Ukraine. Project "Identification of the panel of biomarkers for glioblastoma personalized therapy " (0117U003595) (scientific supervisor Areshkov P.)

Projects of State Fund for Fundamental Researches:

  • 2016 Grant of the President of Ukraine to support scientific research of young scientists. Project "Identification of molecular markers for early diagnosis of tumors and characterization of mechanisms of increased expression of the gene of chitinase 3 of a similar protein 1" (0116U006539) (scientific supervisor Balynska O.)
  • 2016-2017 Joint research projects between the State Fund for Fundamental Research and the Belarusian Republican Fund for Fundamental Research "Loss of heterozygosity and methylation of immune suppressor genes in kidney and bladder carcinomas" (0116U007719 & 0117U002802) (scientific supervisor Skrypkina I.)

Projects of Ministry of Education and Science of Ukraine:

  • 2015, 2016 Ukrainian-Austrian Research Project "Investigation of mechanisms of the formation of chemoresistance of glial tumors and development of approaches to their complex therapy" (0115U005604) (scientific supervisors Dmytrenko V., Avdeev S.)
  • 2016, 2017 Ukrainian-Romanian research project "Identification of optimal combinations of targeted drugs capable of simultaneously blocking proliferative and invasive pathways in glial tumors" (0116U006758 & 0117U003252) ) (scientific supervisors Avdeev S., Areshkov P.)

Collaboration:

with Ukrainian organizations:

  • State Institution Institute of Urology of NAMS of Ukraine (Kyiv)
  • Institute of Cell Therapy (Kyiv)
  • National Cancer Institute, Ministry of Health of Ukraine (Kyiv)
  • State Institution Institute of Neurosurgery named after A. P. Romodanov of NAMS of Ukraine (Kyiv)
  • O. V. Palladin Institute of Biochemistry, NASU (Kyiv)
  • Bogomoletz Institute of Physiology, NASU (Kyiv)
  • Educational and Scientific Centre "Institute of Biology and Medicine" of Taras Shevchenko National University of Kyiv
  • R. E. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology, NASU (Kyiv)
  • The V. N. Karazin Kharkiv National University (Kharkiv)
  • Danylo Halytsky Lviv National Medical University (Lviv)
  • Institute of Cell Biology NASU (Lviv)
  • Clinic-Diagnostic Center "Pharmbioest" (Rubezhne, Lugansk region)

with foreign organizations:

  • Institute of Genetics and Cytology of the National Academy of Sciences of Belarus (Minsk, Belarus)
  • Belarusian State University (Minsk, Belarus)
  • Moffitt Cancer Center (Tampa, Florida).
  • Institute of Gene Biology, RAS (Moscow, Russia)
  • Serbsky State Scientific Center for Social and Forensic Psychiatry (Moscow, Russia)
  • Institute of Chemical Biology and Fundamental Medicine, SB RAS (Novosibirsk, Russia)
  • Institute Gustave-Roussy (Paris, France)
  • Service de Neurologie Mazarin, INSERM U 711 (Paris, France)

    Selected publications:

    1. Svitina H., Skrypkina I. , Areshkov P., Kyryk V., Bukreieva T., Klymenko P., L. Garmanchuk, Lobintseva G., Shablii V. Transplantation of placenta‑derived multipotent cells in rats with dimethylhydrazine‑induced colon cancer decreases survival rate// Oncology Letters, Published online on: February 8, 2018, https://doi.org/10.3892/ol.2018.7996
    2. Stepanenko, and Heng H.H.. Transient and stable vector transfection: Pitfalls, off-target effects, artifacts. Mutation Research. 2017. Vol. 773: 91-103
    3. Svitina H., Kyryk V., Skrypkina I., Kuchma M., Bukreieva T., Areshkov P., Shablii Y., Denis Y., Klymenko P., Garmanchuk L., Ostapchenko L., Lobintseva G., and Shablii V. Placenta-derived multipotent cells have no effect on the size and number of DMH-induced colon tumors in rats. Exp Ther Med. 2017. Vol. 14, N. 3. P. 2135-47.
    4. Skrypkina I., Tsyba L., Onyshchenko K., Morderer D., Kashparova O., Nikolaienko O., Panasenko G., Vozianov S., Romanenko A., Rynditch A. Concentration and methylation of cell-free DNA from blood plasma as diagnostic markers of renal cancer. Disease Markers, 2016, 2016:3693096. DOI: 10.1155/2016/3693096
    5. Stepanenko A.A., Andreieva S.V., Korets K.V., Mykytenko D.O., Baklaushev V.P., Chekhonin V.P., Dmitrenko V.V. mTOR inhibitor temsirolimus and MEK1/2 inhibitor U0126 promote chromosomal instability and cell type-dependent phenotype changes of glioblastoma cells. Gene. 2016, 579 (1):58-68.
    6. Stepanenko A.A. Andreieva S.V. Korets K.V. Mykytenko D.O. Baklaushev V.P. Huleyuk N.L. Kovalova O.A. Kotsarenko K.V. Chekhonin V.P. Vassetzky Y.S. Avdieiev S.S. Dmitrenko V.V. Temozolomide promotes genomic and phenotypic changes in glioblastoma cells. Cancer Cell Int., 36, doi: 10.1186/s12935-016-0311-8.
    7. Dergai M., Iershov A., Novokhatska O., Pankivskyi S., Rynditch A. Evolutionary changes on the way to clathrin-mediated endocytosis in animals. Genome Biol. Evol. 2016, 8(3):588-606.
    8. Finiuk N. S., Senkiv J. V., Riabtseva A. O., Mitina N. Y., Molochii N. I., Kitsera M. O., Avdieiev S. S., Zaichenko O. S., R. S. Stoika. Modulation of temozolomide action towards rat and human glioblastoma cells in vitro by its combination with doxorubicin and immobilization with nanoscale polymeric carrier. Ukr.Biochem. J.2016, 88 (special issue): 87-98.
    9. Skrypkina I. Ya., Onyshchenko K.V., Kashparova O.., Rynditch A. V., Grygorenko V.V., Pereta L.V., Vikarchuk M.V., Banas O.O. Detection of the methylation status of genes LRRC3B, RASSF1A, APC of the cell-free and tumor DNA patients with renal cell carcinoma. Health of men / 2015, Vol. 53 (2): 166-170
    10. Stepanenko A.A., Dmitrenko V.V. HEK293 in cell biology and cancer research: phenotype, karyotype, tumorigenicity, and stress-induced genome-phenotype evolution. Gene. 2015;569(2):182-190.
    11. Stepanenko A.A., Baklaushev V.P., Vassetzky Y.S., Dmitrenko V.V. Cisplatin treatment of C6 rat glioma in vivo did not influence copy number alterations and growth pattern of tumor-derived resistant cells. Biopolym. Cell. 2015; 31(3):209217.
    12. Avdieiev S., Gera L., Hodges R.S., Dmitrenko V.V. Glioma-associated protein CHI3L2 suppresses cells viability and induces G1/S transition arrest. Biopolym. Cell. 2015;31(4):316-320.
    13. Stepanenko A, Andreieva S, Korets K, Mykytenko D, Huleyuk N, Vassetzky Y, Kavsan V. Step-wise and punctuated genome evolution drive phenotype changes of tumor cells. Mutation Res. (Mutation Research-Fundamental and Molecular Mechanisms of Mutagenesis). 2015;771:56-69.
    14. Morderer D. Ye., Rymarenko O. V., Skrypkina I. Ya., Rynditch A. V. Ca2+ does not affect the binding properties of ITSN1 EH domains. Biopolym. Cell. 2014;30(6):481-484.
    15. Avdieiev S, Gera L, Havrylyuk D, et al. Bradykinin antagonists and thiazoli-dinone derivatives as new potential anti-cancer compounds. Bioorg. & Med. Chem. 2014;22:3815-3823.
    16. Stepanenko AA, Kavsan VM. Karyotypically distinct U251, U373, and SNB19 glioma cell lines are of the same origin but have different drug treatment sensitivities. Gene. 2014;540:263-265.
    17. Morderer DYe, Nikolaienko OV, Skrypkina IYa, et al. Ca/calmodulin-dependent phosphorylation of endocytic scaffold ITSN1. Biopolymers and Cell. 2014;30(1):74-76.
    18. Kavsan VM, Kulagova TA, Kuznetsova TA, et al. Structure and function of oncogene-transfected immortal cells. Biopolym. Cell. 2014;30(1):25-28.
    19. Avdieiev SS, Gera L, Hodges R, et al. Growth suppression activity of bradykinin antagonists in glioma cells. Biopolym. Cell. 2014;30(1):77-79.
    20. Morderer D. Ye., Nikolaienko O. V., Skrypkina I. Ya., Rymarenko O. V., Kropyvko S. V., Tsyba L. O., Rynditch A. V. Ca/calmodulin-dependent phosphorylation of endocytic scaffold ITSN1. Biopolym. Cell. 2014;30(1):74-76
    21. Novokhatska O., Dergai M., Tsyba L., Skrypkina I., Filonenko V., Moreau J., Rynditch A. Adaptor Proteins Intersectin 1 and 2 Bind Similar Proline-Rich Ligands but Are Differentially Recognized by SH2 Domain-Containing Proteins. PLoS One, 2013, Vol.8 (7):e70546.
    22. Tsyba L. O., Dergai M. V., Skrypkina I. Ya., Nikolaienko O. V., Dergai O. V., Kropyvko S. V., Novokhatska O. V., Morderer D. Ye., Gryaznova T. A., Gubar O. S., Rynditch A. V. ITSN protein family: regulation of diversity, role in signalling and pathology. Biopolym. Cell. 2013;29(3):244-251.
    23. Dergai O, Dergai M, Skrypkina I, Matskova L, Tsyba L, Gudkova D, Rynditch A The LMP2A protein of Epstein-Barr virus regulates phosphorylation of ITSN1 and Shb adaptors by tyrosine kinases. Cell Signal. 2013, Vol.25(1), P:33-40
    24. Stepanenko AA, Vassetzky YS, Kavsan VM. Antagonistic functional duality of cancer genes. Gene. 2013; 529: 199-207. doi: 10.1016/j.gene.2013.07.047
    25. Stepanenko AA, Kavsan VM. Cancer genes and chromosome instability. Oncogene and Cancer From Bench to Clinic, InTech Publisher, 2013; 151-182.
    26. Dmitrenko VV, Avdieiev SS, Areshkov PO, et al. From reverse transcription to human brain tumors Biopolym. Cell. 2013; 29(3):221-233 doi: 10.7124/bc.00081C
    27. Skrypkina .Ya., Kondratov .G., Tsyba L.O., Panasenko G.V., Nikolaienko .V., Romanenko .., Kolesnyk O.O., Morderer D.Ye., Nekrasov K.A., shuba V.., Vozianov S.O., Shchepotin I.B., Rynditch .V. Detection of cell-free DNA and gene-specific methylation in blood plasma of patients with renal and colon cancer. 2012, Science and Innovation, 2012, Vol.8(6), P:60-66.
    28. Morderer D, Nikolaienko O, Skrypkina I, Cherkas V, Tsyba L, Belan P, Rynditch A. Endocytic adaptor protein intersectin 1 forms a complex with microtubule stabilizer STOP in neurons. Gene. 2012, Vol.505(2), P:360-364.
    29. Stepanenko AA, Kavsan VM. Evolutionary karyotypic theory of cancer versus conventional cancer gene mutation theory. Biopolym. Cell. 2012; 28(4):267280. doi:10.7124/bc.000059
    30. Areshkov PO, Avdieiev SS, Balynska OV, LeRoith D, Kavsan VM. Two closely related human members of chitinaselike family, CHI3L1 and CHI3L2, activate ERK1/2 in 293 and U372 cells but have the different influence on cell proliferation. Int. J. Biol. Sci. 2012; 8: 39-48.
    31. Dergai M, Skrypkina I, Dergai O, Tsyba L, Novokhatska O, Filonenko V, Drobot L, Rynditch A. Identification and characterization of a novel mammalian isoform of the endocytic adaptor ITSN1.Gene. 2011, Vol.485, 120-129
    32. Kavsan VM, Baklaushev VP, Balynska OV, et al. Gene encoding chitinase 3like 1 protein (CHI3L1) is a putative oncogene. Int. J. Biomed. Sci. 2011; 7: 230237.
    33. Kavsan VM, Iershov AV, Balynska OV. Immortalized cells and one oncogene in malignant transformation: old insights on new explanation. BMC Cell Biol. 2011; 12:23. doi: 10.1186/1471-2121-12-23
    34. Tsyba L., Nikolaienko O., Dergai O., Dergai M., Novokhatska O., Skrypkina I. and Rynditch, A. (2011) Intersectin multidomain adaptor proteins: Regulation of functional diversity. Gene, Vol.473, 67-75.