Obtaining a stable anti-tumor phenotype of THP-1 cells for cancer treatment

¹ National Medical Research Center for Obstetrics, Gynecology and Perinatology named after Academician V.I.Kulakov of Ministry of Healthcare of Russian Federation
² Research Institute of Molecular and Cellular Medicine, Peoples' Friendship University of Russia
³ ederal State Budgetary Institution "National Medical Research Center for Obstetrics, Gynecology and Perinatology named after Academician V.I. Kulakov"
⁴ Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Pirogov Russian National Research Medical University
⁵ N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia, 115478 Moscow, Russian Federation
⁶ Sirius University of Science and Technology
⁷ Avtsyn Research Institute of Human Morphology of Federal state budgetary scientific institution "Petrovsky National Research Centre of Surgery"

Academic Editor: Jurg Bahler

Published: 12 April 2024 by MDPI in The 3rd International Electronic Conference on Biomolecules session Molecular Mechanisms in Cellular Processes

Abstract:

Introduction. Macrophages are cells of the innate immune system responsible for phagocytosis and antigen presentation. According to the modern concept of binary polarization, pro- (M1) and anti-inflammatory macrophages (M2) are distinguished. It is believed that M2 macrophages, by inhibiting the T-cell anti-tumor immune response and participating in tumor angiogenesis, can lead to tumor progression. On the other hand, M1 macrophages exert an anti-tumor effect through their participation in antibody-dependent cell-mediated cytotoxicity. The goal of this work was to develop a new method of cellular anti-tumor therapy based on M1 macrophages.

Methods. After analysis of M1 in vitro polarization transcriptomic data, the BIN1, Ido1, Asic1, Sosc3, Socs1, Irf4, Irf5 genes were selected for knockout. To form a stable pro-inflammatory phenotype, genetic modification of THP-1 cells based on the CRISPR/Cas system was carried out using lentiviral transduction. After transduction, infected THP-1 cells were sorted and their anti-tumor activity was tested in vivo and in vitro using various methods.

Results. Evaluation of knockout cells by real-time PCR showed a significant decrease in the expression level of the Asic1 gene. The use of infected knockout cells in a mouse model of breast cancer showed a decrease in tumor growth relative to control at one time point. Evaluation of Ki67 using immunohistochemistry analysis showed a significant decrease in Ki67+ cells in tumors treated with THP1 knockout cells. When analyzing surface markers of macrophages using flow cytometry in the group treated with knockout cells, we revealed a significant increase in marker CD45, monocyte marker CD11b, macrophage marker CD68 and the pro-inflammatory macrophage marker CD86.

Conclusions. The results obtained indicate the successful genetic modification of THP-1 to obtain a pro-inflammatory phenotype, and also suggest the effectiveness of M1-based cell therapy for tumor diseases. This work was supported by a grant from the Russian Science Foundation [grant number 22-15-00241].

Keywords: macrophages; monocytes; M1; tumor; lentiviral transduction;