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Establishment of a FDC-P1 murine cell line with human KIT N822K gene overexpression

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1. Title Title of document Establishment of a FDC-P1 murine cell line with human KIT N822K gene overexpression
2. Creator Author's name, affiliation, country Elmira R. Vagapova; Engelhardt Institute of Molecular Biology, Russian Academy of Sciences; Россия
2. Creator Author's name, affiliation, country T. D. Lebedev; Engelhardt Institute of Molecular Biology, Russian Academy of Sciences; Россия
2. Creator Author's name, affiliation, country V. I. Popenko; Engelhardt Institute of Molecular Biology, Russian Academy of Sciences; Россия
2. Creator Author's name, affiliation, country O. G. Leonova; Engelhardt Institute of Molecular Biology, Russian Academy of Sciences; Россия
2. Creator Author's name, affiliation, country P. V. Spirin; Engelhardt Institute of Molecular Biology, Russian Academy of Sciences; Россия
2. Creator Author's name, affiliation, country V. S. Prasolov; Engelhardt Institute of Molecular Biology, Russian Academy of Sciences; Россия
3. Subject Discipline(s)
3. Subject Keyword(s) receptor tyrosine kinase KIT; FDC-P1; acute myeloid leukemia (AML); KIT N822K; stromal cells
4. Description Abstract

The mechanism of resistance of leukemia cells to chemotherapeutic drugs remains poorly understood. New model systems for studying the processes of malignant transformation of hematopoietic cells are needed. Based on cytokine-dependent murine acute myeloid leukemia (AML) FDC-P1 cells, we generated a new cell line with ectopic expression of the KIT gene encoding mutant human receptor tyrosine kinase (N822K). We investigated the role played by overexpression of the mutant KIT in the survival of leukemia cells and their sensitivity to therapeutic drugs. We also generated a co-culture system consisting of FDC-P1 murine leukemia cells and a HS-5 human stromal cell line. Our data can be used for a further comprehensive analysis of the role of KIT N822K mutation in the cellular response to anti-leukemic drugs, growth factors, and cytokines. These data are of interest in the development of new effective therapeutic approaches to the treatment of acute leukemia.

5. Publisher Organizing agency, location Acta Naturae Ltd
6. Contributor Sponsor(s) Russian Foundation for Basic Research (18-29-09151)
Russian Foundation for Basic Research (17-04-01555)
7. Date (DD-MM-YYYY) 16.04.2020
8. Type Status & genre Peer-reviewed Article
8. Type Type Research Article
9. Format File format
10. Identifier Uniform Resource Identifier https://actanaturae.ru/2075-8251/article/view/10938
10. Identifier Digital Object Identifier (DOI) 10.32607/actanaturae.10938
11. Source Title; vol., no. (year) Acta Naturae; Vol 12, No 1 (2020)
12. Language English=en ru
13. Relation Supp. Files Fig. 1. Schematic representation of the vector for the expression of the human KIT N822K gene (A); isolation of the population of cells with high fluorescence intensity in the FITC channel (highlighted with an ellipse) by cell sorting (B), the population of non-transduced FDC-P1 cells is shown in gray, cells transduced with the vector carrying KIT N822K are shown in green; surface KIT receptor expression in the control cells and cells showing overexpression of the mutant human KITN 822K (C); cells were treated with PerCP5.5-conjugated monoclonal antibodies to human KIT and analyzed by flow cytometry: untreated cells are shown in gray, cells treated with human KIT-specific antibodies are shown in pink; the expression level of the murine KIT (mus) and human KIT (hum) genes in FDC-P1 cells with overexpression of KIT N822K according to the qPCR data (D); immunocytochemical evaluation of the expression of the human KIT receptor gene (pink) in FDC-P1 cells on a confocal microscope (E); nuclei are stained with DAPI (green) (E) (588KB) doi: 10.32607/20758251-2020-12-1-51-55-268
Fig. 2. Growth curve of FDC-P1 cells in the presence of IL-3 (solid line) and in a reduced IL-3 content (dotted line) (A); the number of FDC-P1 cells treated with recombinant SCF for 4 days per milliliter (B); the number of FDC-P1 cells as a percentage relative to the untreated cells on day 3 after addition of imatinib (C) and cytarabine (D) drugs. *p < 0.05 (687KB) doi: 10.32607/20758251-2020-12-1-51-55-269
Fig. 3. The number of FDC-P1 cells cultured in the absence (green) and presence of HS-5 human stromal cells (pink) for 3 (A) and 5 (B) days; images of FDC-P1 cells co-cultured with HS-5 cells: control (C) and N822K (D) cells. Longitudinal stromal cells adhere to the plate bottom, while round FDC-P1 cells remain in suspension (316KB) doi: 10.32607/20758251-2020-12-1-51-55-270
14. Coverage Geo-spatial location, chronological period, research sample (gender, age, etc.)
15. Rights Copyright and permissions Copyright (c) 2020 Vagapova E.R., Lebedev T.D., Popenko V.I., Leonova O.G., Spirin P.V., Prasolov V.S.
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