ISSN Print 2500–1094
ISSN Online 2542–1204
Bulletin of RSMU
BIOMEDICAL JOURNAL OF PIROGOV UNIVERSITY (MOSCOW, RUSSIA)
ORIGINAL RESEARCH
Immunogenicity of full-length and multi-epitope mRNA vaccines for M. Tuberculosis as demonstrated by the intensity of T-cell response: a comparative study in mice
About authors
1 Translational Medicine Research Center, Sirius University of Science and Technology (Autonomous Non-Commercial Higher Education Organization), Sirius, Russia
2 Pavlov First Saint Petersburg State Medical University, St. Petersburg, Russia
3 Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia
Correspondence should be addressed: Vasily V. Reshetnikov
Olympiyskiy prospekt, 1, Sochi, 354340, Russia; ur.hepsuitnalat@vv.vokintehser
About paper
Funding: the study was supported by the Ministry of Science and Higher Education of the Russian Federation (agreement № 075-10-2021-113, unique project identifier RF----193021X0001).
Acknowledgements: the authors express their gratitude to the staff of the Sirius University: Terenin IM for in vitro transcription, Zaborova OV for the formulation of mRNA into lipid nanoparticles, Gavrilyak MA and Golovin EV for purification and characterization of the Rv3875 protein; Shevyrev DV for setting up the intracellular staining experiment and Sitikova VA for assistance in the context of the animal experiment.
Author contribution: Vasileva OO, Tereschenko VP — work with the cells, data analysis, article authoring, figures authoring; Krapivin BN, Pateev II, Rybtsov SA — work with the cells, animal experiment, data analysis; Muslimov AR, Kukushkin IS — design of structures, cloning; Ivanov RA, Reshetnikov VV — text editing, data analysis, project coordination.
Compliance with the ethical standards: animal study was approved by the Ethics Committee of the Institute of Cytology and Genetics (2022); carried out in accordance with the laboratory animals care guidelines (2010), Directive 2010/63/EU of the European parliament and of the Council on the protection of animals used for scientific purposes (2010), Good Laboratory Practice guidelines (2016).
Received: 2023-04-24
Accepted: 2023-06-15
Published online: 2023-06-29
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Fig. 1. Experiment design. A. Schematic representation of composition of the mRNA-LNP vaccine and other drugs. B. Design of the experiment in C57BL6/J mice. The two different variants of mRNA vaccines — mRNA Rv3875 (582 nucleotides) or mRNA mEpitope (699 nucleotides), 50 µg of RNA each, — were administered intramuscularly. In addition, mice were immunized with full-length Rv3875 protein (25 mg) adjuvanted with mRNA-free lipid nanoparticles in an amount equivalent to (±10%) the number of particles in the RNA vaccine groups. Besides the test groups, there were two control groups in the experiment, first of which received lipid nanoparticles without mRNA, second — a phosphate buffer. Each group included three animals. Second immunization (same doses) took place 4 weeks after the first one. The mice were killed 4 weeks after second immunization and their spleen isolated. We used the ELISpot assay to count splenocytes secreting IFNγ in response to the Rv3875 tuberculosis protein post vaccination, and thus measured the level of the resulting T-cell response
Fig. 2. Evaluation of the T-cell response of splenocytes of the immunized mice. A. ELISpot results. Number of cells secreting IFNγ in response to DC2.4 stimulation, activated by the Rv3875 protein. The data are presented as mean ± error of mean. Three animals per group. * — p < 0.05 compared with the PBS group; # — p < 0.05 compared with the LNP group; & — p < 0.05 compared with the Rv3875 + LNP group. B. Representative data on CD3+ T lymphocytes (CD8+ and CD8– ) producing IFNγ (intracellular staining). The figures show the percentage of cells from the total pool of CD3+ CD8+ and CD3+ CD8– T-lymphocytes that produce IFNγ
