2023  1,500
2022  1,200
2021  1,540
2020  1,374
2019  1,023
2018  0,932
2017  0,977
2016  0,799
2015  0,662
2014  0,740
2013  0,739
2012  0,637
2011  0,658
2010  0,654
2009  0,570
2008  0,849
2007  0,805
2006  0,330
2005  0,435
2004  0,623
2003  0,567
2002  0,641
2001  0,490
2000  0,477
1999  0,762
1998  0,785
1997  0,507
1996  0,518
1995  0,502
Vol 58(2024) N 1 p. 147-156; DOI 10.1134/S0026893324010102 Full Text

N.I. Ponomareva1,2,3*, S.A. Brezgin1,2, A.P. Kostyusheva1, O.V. Slatinskaya4, E.O. Bayurova5, I.V. Gordeychuk5, G.V. Maksimov4, D.V. Sokolova7, G. Babaeva7, I.I. Khan7, V.S. Pokrovsky7, A.S. Lukashev1, V.P. Chulanov6, D.S. Kostyushev1,2

Stochastic Packaging of Cas Proteins into Exosomes

1Martsinovsky Institute of Medical Parasitology, Tropical and Vector-Borne Diseases, First Moscow State Medical University (Sechenov University), Moscow, 119991 Russia
2Sirius University of Science and Technology, Sochi, 354340 Russia
3Department of Pharmaceutical and Toxicological Chemistry, First Moscow State Medical University (Sechenov University), Moscow, 119146 Russia
4Faculty of Biology, Moscow State University, Moscow, 119991 Russia
5Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products, Russian Academy of Sciences, Moscow, 108819 Russia
6National Medical Research Center of Tuberculosis and Infectious Diseases, Ministry of Health, Moscow, 127994 Russia
7N.N. Blokhin National Medical Research Center of Oncology, Moscow, 115478 Russia

Received - 2023-06-17; Revised - 2023-08-09; Accepted - 2023-08-23

CRISPR/Cas systems are perspective molecular tools for targeted manipulation with genetic materials, such as gene editing, regulation of gene transcription, modification of epigenome etc. While CRISPR/Cas systems proved to be highly effective for correcting genetic disorders and treating infectious diseases and cancers in experimental settings, clinical translation of these results is hampered by the lack of efficient CRISPR/Cas delivery vehicles. Modern synthetic nanovehicles based on organic and inorganic polymers have many disadvantages, including toxicity issues, the lack of targeted delivery, and complex and expensive production pipelines. In turn, exosomes are secreted biological nanoparticles that exhibit high biocompatibility, physico-chemical stability, and the ability to cross biological barriers. Early clinical trials found no toxicity associated with exosome injections. In the recent years, exosomes have been considered as perspective delivery vehicles for CRISPR/Cas systems in vivo. The aim of this study was to analyze the efficacy of CRISPR/Cas stochastic packaging into exosomes for several human cell lines. Here, we show that Cas9 protein is effectively localized into the compartment of intracellular exosome biogenesis, but stochastic packaging of Cas9 into exosomes turns to be very low (~1%). As such, stochastic packaging of Cas9 protein is very ineffective and cannot be used for gene editing purposes. Developing novel tools and technologies for loading CRISPR/Cas systems into exosomes is needed.

delivery, nanoparticles, CRISPR/Cas, extracellular vesicles, exosomes, gene editing