Comprehensive mRNA-sequencing-based characterization of three HEK-293 cell lines during an rAAV production process for gene therapy applications.
HEK-293 cells
gene therapy
rAAV
transcriptomics
transient transfection
virus production
Journal
Biotechnology journal
ISSN: 1860-7314
Titre abrégé: Biotechnol J
Pays: Germany
ID NLM: 101265833
Informations de publication
Date de publication:
Aug 2023
Aug 2023
Historique:
revised:
20
04
2023
received:
11
10
2022
accepted:
10
05
2023
medline:
17
8
2023
pubmed:
16
5
2023
entrez:
16
5
2023
Statut:
ppublish
Résumé
Human embryonal kidney cells (HEK-293) are the most common host cells used for transient recombinant adeno-associated virus (rAAV) production in pharmaceutical industry. To better cover the expected gene therapy product demands in the future, different traditional strategies such as cell line sub-cloning and/or addition of chemical substances to the fermentation media have been used to maximize titers and improve product quality. A more effective and advanced approach to boost yield can be envisaged by characterizing the transcriptome of different HEK-293 cell line pedigrees with distinct rAAV productivity patterns to subsequently identify potential gene targets for cell engineering. In this work, the mRNA expression profile of three HEK-293 cell lines, resulting in various yields during a fermentation batch process for rAAV production, was investigated to gain basic insight into cell variability and eventually to identify genes that correlate with productivity. Mock runs using only transfection reagents were performed in parallel as a control. It finds significant differences in gene regulatory behaviors between the three cell lines at different growth and production stages. The evaluation of these transcriptomics profiles combined with collected in-process control parameters and titers shed some light on potential cell engineering targets to maximize transient production of rAAV in HEK-293 cells.
Identifiants
pubmed: 37191240
doi: 10.1002/biot.202200513
doi:
Substances chimiques
RNA, Messenger
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e2200513Informations de copyright
© 2023 The Authors. Biotechnology Journal published by Wiley-VCH GmbH.
Références
Zhao, Z., Anselmo, A. C., & Mitragotri, S. (2022). Viral vector-based gene therapies in the clinic. Bioengineering & Translational Medicine, 7, e10258.
Sarvari, R., Nouri, M., Agbolaghi, S., Roshangar, L., Sadrhaghighi, A., Seifalian, A. M., & Keyhanvar, P. (2020). A summary on non-viral systems for gene delivery based on natural and synthetic polymers. International Journal of Polymeric Materials and Polymeric Biomaterials, 71, 246-65.
Shahryari, A., Saghaeian Jazi, M., Mohammadi, S., Razavi Nikoo, H., Nazari, Z., Hosseini, E. S., Burtscher, I., Mowla, S. J., & Lickert, H. (2019). Development and clinical translation of approved gene therapy products for genetic disorders. Front Genet, 10, 868.
Kimberly, D. (2008). Toward development of artificial viruses for gene therapy: A comparative evaluation of viral and non-viral transfection. Biotechnology Progress, 24, 871-83.
Patil, G., Lin, L., Dang, T., Zhang, J., & Qadir, Q. (2019). The development of functional non-viral vectors for gene delivery. International Journal of Molecular Sciences, 20(21), 5491.
Ramamoorth, M. (2015). Non viral vectors in gene therapy- an overview. Journal of Clinical and Diagnostic Research, 9, GE01-6.
Ren, S., Wang, M., Wang, C., Wang, Y., Sun, C., Zeng, Z., Cui, H., & Zhao, X. (2021). Application of non-viral vectors in drug delivery and gene therapy. Polymers (Basel), 13(19), 3307.
Graham, F. L., Smiley, J., Russell, W. C., & Nairn, R. (1977). Characteristics of a human cell line transformed by DNA from human adenovirus type 5. Journal of General Virology, 36, 59-74.
Acosta-Alvear, D., Zhou, Y., Blais, A., Tsikitis, M., Lents, N. H., Arias, C., Lennon, C. J., Kluger, Y., & Dynlacht, B. D. (2007). XBP1 controls diverse cell type- and condition-specific transcriptional regulatory networks. Molecular Cell, 27, 53-66.
Oxgene. 2023. The role of HEK293 cells in gene therapy. New-Medical life sciences. Retrieved on March 15, 2023 from https://www.news-medical.net/whitepaper/20200520/The-Role-of-HEK293-Cells-in-Gene-Therapy.aspx
Fiszer-Kierzkowska, A., & Vydra, N. (2011). Liposome-based DNA carriers may induce cellular stress response and change gene expression pattern in transfected cells. BMC Molecular Biology, 12(1), 10.1186/1471-2199-12-27
Jacobsen, L., Calvin, S., & Lobenhofer, E. (2009). Transcriptional effects of transfection: The potential for misinterpretation of gene expression data generated from transiently transfected cells. Biotechniques, 47, 617-24.
Lin, Y. (2014). Genome dynamics of the human embryonic kidney 293 lineage in response to cell biology manipulations. Nature Communications, 5(1), 4767. https://doi.org/10.1038/ncomms5767
Bushnell, B. (2013). "BBMap." In, BBMap: Short read aligner for DNA and RNA-seq data. Capable of handling arbitrarily large genomes with millions of scaffolds. Handles Illumina, PacBio, 454, and other reads; very high sensitivity and tolerant of errors and numerous large indels.
Andrews, S. (2010). "FASTQC." In, FastQC aims to provide a simple way to do some quality control checks on raw sequence data coming from high throughput sequencing pipelines. It provides a modular set of analyses which you can use to give a quick impression of whether your data has any problems of which you should be aware before doing any further analysis.
Yates, A. D., Achuthan, P., Akanni, W., Allen, J., Allen, J., Alvarez-Jarreta, J., Amode, M. R., Armean, I. M., Azov, A. G., Bennett, R., Bhai, J., Billis, K., Boddu, S., Marugán, J. C., Cummins, C., Davidson, C., Dodiya, K., Fatima, R., Gall, A., … Flicek, P. (2020). Ensembl 2020. Nucleic Acids Research, 48, D682-D88.
Dobin, A., Davis, C. A., Schlesinger, F., Drenkow, J., Zaleski, C., Jha, S., Batut, P., Chaisson, M., & Gingeras, T. R. (2013). STAR: Ultrafast universal RNA-seq aligner, Bioinformatics, 29, 15-21.
Putri, G H., Anders, S., Pyl, P. T., Pimanda, J E., Zanini, F., & Boeva, V. (2022). Analysing high-throughput sequencing data in Python with HTSeq 2.0. Bioinformatics, 38, 2943-45.
Mootha, V. K., Lindgren, C. M., Eriksson, K.-F., Subramanian, A., Sihag, S., Lehar, J., Puigserver, P., Carlsson, E., Ridderstråle, M., Laurila, E., Houstis, N., Daly, M. J., Patterson, N., Mesirov, J. P., Golub, T. R., Tamayo, P., Spiegelman, B., Lander, E. S., Hirschhorn, J. N., … Groop, L. C. (2003). PGC-1α-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nature Genetics, 34(3), 267-273.
Subramaniana, A. (2005). Gene set enrichment analysis: A knowledge-based approach for interpreting genome-wide expression profiles. Proceedings of the National Academy of Sciences, 102(43), 15545-15550.
Schneider, M., & Marison, I. (1996). The importance of ammonia in mammalian cell culture. Journal of Biotechnol, 46, 161-85.
Tsang, S. M., Oliemuller, E., & Howard, B. A. (2020). Regulatory roles for SOX11 in development, stem cells and cancer. Seminars in Cancer Biology, 67, 3-11.
Decaesteker, B., Louwagie, A., Loontiens, S., De Vloed, F., Roels, J., Vanhauwaert, S., De Brouwer, S., Sanders, E., Denecker, G., D'haene, E., Van Haver, S., Van Loocke, W., Van Dorpe, J., Creytens, D., Van Roy, N., Pieters, T., Van Neste, C., Fischer, M., Van Vlierberghe, P., … Speleman, F. (2020). SOX11 is a lineage-dependency factor and master epigenetic regulator in neuroblastoma. bioRxiv, 2020, S. 2020.08. 21.261131.
Dabbaghi, M., Kazemi Oskuee, R., Hashemi, K., & Afkhami Goli, A. (2018). Evaluating polyethyleneimine/DNA nanoparticles-mediated damage to cellular organelles using endoplasmic reticulum stress profile. Artif Cells Nanomed Biotechnol, 46, 192-99.
Roman, R., Farras, M., Camps, M., Martinez-Monge, I., Comas, P., Martinez-Espelt, M., Lecina, M., Casablancas, A., & Cairo, J. J. (2018). Effect of continuous feeding of CO2 and pH in cell concentration and product titers in hIFNγ producing HEK293 cells: Induced metabolic shift for concomitant consumption of glucose and lactate. Journal of Biotechnology, 287, 68-73.
Lavorgna, M., Russo, C., D'Abrosca, B., Parrella, A., & Isidori, M. (2016). Toxicity and genotoxicity of the quaternary ammonium compound benzalkonium chloride (BAC) using Daphnia magna and Ceriodaphnia dubia as model systems. Environmental Pollution, 210, 34-9.
Moghimi, S. M., Symonds, P., Murray, J. C., Hunter, A. C., Debska, G., & Szewczyk, A. (2005). A two-stage poly(ethylenimine)-mediated cytotoxicity: Implications for gene transfer/therapy. Molecular Therapy, 11, 990-5.
Parhamifar, L., Larsen, A. K., Hunter, A. C., Andresen, T. L., & Moghimi, S. M. (2010). Polycation cytotoxicity: A delicate matter for nucleic acid therapy-focus on polyethylenimine. Soft Matter, 6, 4001-09.
Unfried, K., Albrecht, C., Klotz, L.-O., Mikecz, A. V., Grether-Beck, S., & Schins, R P. F. (2009). Cellular responses to nanoparticles: Target structures and mechanisms. Nanotoxicology, 1, 52-71.
Yoshida, H., Matsui, T., Yamamoto, A., Okada, T., & Mori, K. (2001). XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell Press, 107, 881-91.
Park, S. M., Kang, T.-I., & So, J.-S. (2021). Roles of XBP1s in transcriptional regulation of target genes. Biomedicines, 9(7), 791.
Formas-Oliveira, A. S., Basilio, J. S., Rodrigues, A. F., & Coroadinha, A. S. (2020). Overexpression of ER protein processing and apoptosis regulator genes in human embryonic kidney 293 cells improves gene therapy vectors production, Biotechnology Journal, 15, e1900562.
Maxfield, F. R., & van Meer, G. (2010). Cholesterol, the central lipid of mammalian cells. Current Opinion in Cell Biology, 22, 422-9.
Sheng, R., Chen, Y., Yung Gee, H., Stec, E., Melowic, H. R., Blatner, N. R., Tun, M. P., Kim, Y., Källberg, M., Fujiwara, T. K., Hye Hong, J., Pyo Kim, K., Lu, H., Kusumi, A., Goo Lee, M., & Cho, W. (2012). Cholesterol modulates cell signaling and protein networking by specifically interacting with PDZ domain-containing scaffold proteins. Nature Communications, 3, 1249.
Maxfield, F. R., & Menon, A. K. (2006). Intracellular sterol transport and distribution. Current Opinion in Cell Biology, 18, 379-85.
Anderson, R. H., Sochacki, K. A., Vuppula, H., Scott, B. L., Bailey, E. M., Schultz, M. M., Kerkvliet, J. G., Taraska, J. W., Hoppe, A. D., & Francis, K. R. (2021). Sterols lower energetic barriers of membrane bending and fission necessary for efficient clathrin-mediated endocytosis. Cell reports, 37, 110008.
Mesmin, B., & Maxfield, F. R. (2009). Intracellular sterol dynamics. Biochimica Et Biophysica Acta, 1791, 636-45.
Xu, F., Rychnovsky, S. D., Belani, J. D., Hobbs, H. H., Cohen, J. C., & Rawson, R. B. (2005). Dual roles for cholesterol in mammalian cells. Biochemistry, 102, 14551-56.
Yeagle, P. L. (1991). Modulation of membrane function by cholesterol. Biochimie, 73, 1303-10.
Zhang, X., Barraza, K. M., & Beauchamp, J. L. (2018). Cholesterol provides nonsacrificial protection of membrane lipids from chemical damage at air-water interface. PNAS, 115, 3255-60.
Pichler, H., & Riezman, H. (2004). Where sterols are required for endocytosis. Biochimica Et Biophysica Acta, 1666, 51-61.
Ouweneel, A. B., Thomas, M. J., & Sorci-Thomas, M. G. (2020). The ins and outs of lipid rafts: Functions in intracellular cholesterol homeostasis, microparticles, and cell membranes. Journal of Lipid Research, 61, 676-86.
Runz, H., Miura, K., Weiss, M., & Pepperkok, R. (2006). Sterols regulate ER-export dynamics of secretory cargo protein ts-O45-G. Embo Journal, 25, 2953-65.
Dunn, J., & Grider, M. H. (2022). Physiology, Adenosine triphosphate. StatPearls.
Alabduladhem, T. O., & Bordoni, B. (2022). Physiology, Krebs cycle. StatPearls.
Chaudhry, R., & Varacallo, M. (2022). Biochemistry, glycolysis. StatPearls.
Deshpande, O. A., & Mohiuddin, S. S. (2022). Biochemistry, oxidative phosphorylation. StatPearls.
Weishaupt, A., & Kadenbach, B. (1992). Selective removal of subunit VIb increases the activity of cytochrome c oxidase. Biochemistry, 31, 11477-81.