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CRISPR/Cas9 mediated genome editing technology to knock out the urokinase receptor gene in neuroblastoma cells


K. D. Rysenkova – Post-graduate Student, Faculty of Medicine, Lomonosov Moscow State University
E. V. Semina – Ph.D. (Biol.), Leading Research Scientist, Federal State Budget Organization National Medical Research Center of Cardiology of the Ministry of Healthcare of the Russian Federation; Senior Research Investigator, Faculty of Medicine, Lomonosov Moscow State University
M. N. Karagyaur – Ph.D. (Biol.), Senior Research Scientist, Institute for Regenerative Medicine, Medical Research and Education Center, Lomonosov Moscow State University
А. А. Shmakova – Student, Faculty of Medicine, Lomonosov Moscow State University
D. T. Dyjkanov – Leading Specialist, Faculty of Medicine, Lomonosov Moscow State University
K. А. Rubina – Dr.Sc. (Biol.), Senior Research Scientist, Faculty of Medicine, Lomonosov Moscow State University
V. А. Tkachuk – Academician RAS, Dr.Sc. (Biol.), Professor, Federal State Budget Organization National Medical Research Center of Cardiology of the Ministry of Healthcare of the Russian Federation; Dean of Fa-culty of Medicine, Lomonosov Moscow State University

Neuroblastoma is considered to be one of the most aggressive pediatric tumors with high invasive potential and risk of me-tastasis. The increased expression of urokinase and its receptor is associated with a negative prognosis of the disease as it promotes tumor proliferation and increases metastatic potential. Nowadays CRISPR/Cas9 technology is widely used for genome editing in mammalian cells. In current study we selected the optimal sequences of single guide RNAs and conditions for the analysis of urokinase receptor gene (uPAR) knockout in linear mouse neuroblastoma cells (Neuro2a) having an unstable genome. To assess the effectiveness of uPAR knockout with CRISPR/Cas9 in Neuro2a clones selected after transfection, the amount of uPAR protein was analyzed by Western blotting. It was found that uPAR knockout decreased uPA-stimulated invasive potential of the cells. Thus, uPAR knockout by CRISPR is a promising therapeutic strategy and it can be considered as a therapeutic approach to suppress migration and invasion of tumor cells.

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May 29, 2020

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