Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells

Strasser, Lisa; Boi, Stefano; Guapo, Felipe; Donohue, Nicholas; Barron, Niall; Rainbow-Fletcher, Alana; Bones, Jonathan

doi:10.3390/ijms222111499

Open AccessArticle

Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells

by

Lisa Strasser

¹,

Stefano Boi

¹

,

Felipe Guapo

¹,

Nicholas Donohue

¹,

Niall Barron

^1,2,

Alana Rainbow-Fletcher

³ and

Jonathan Bones

^1,2,*

¹

NIBRT—National Institute for Bioprocessing Research and Training, Foster Avenue, Blackrock, A94 X099 Dublin, Ireland

²

School of Chemical and Bioprocess Engineering, University College Dublin, Belfield, D04 V1W8 Dublin, Ireland

³

Pharmaron, 12 Estuary Banks, Speke, Liverpool L24 8RB, UK

^*

Author to whom correspondence should be addressed.

Int. J. Mol. Sci. 2021, 22(21), 11499; https://doi.org/10.3390/ijms222111499

Submission received: 29 August 2021 / Revised: 18 October 2021 / Accepted: 20 October 2021 / Published: 25 October 2021

(This article belongs to the Special Issue MS-Based Protein Specific Analysis)

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Abstract

Adeno-associated viral (AAV) vectors are widely used for gene therapy, providing treatment for diseases caused by absent or defective genes. Despite the success of gene therapy, AAV manufacturing is still challenging, with production yields being limited. With increased patient demand, improvements in host cell productivity through various engineering strategies will be necessary. Here, we study the host cell proteome of AAV5-producing HEK293 cells using reversed phase nano-liquid chromatography and tandem mass spectrometry (RPLC-MS/MS). Relative label-free quantitation (LFQ) was performed, allowing a comparison of transfected vs. untransfected cells. Gene ontology enrichment and pathway analysis revealed differential expression of proteins involved in fundamental cellular processes such as metabolism, proliferation, and cell death. Furthermore, changes in expression of proteins involved in endocytosis and lysosomal degradation were observed. Our data provides highly valuable insights into cellular mechanisms involved during recombinant AAV production by HEK293 cells, thus potentially enabling further improvements of gene therapy product manufacturing.

Keywords: proteomics; LC-MS/MS; SP3; label-free quantitation; HEK293 cells; adeno-associated virus (AAV); gene therapy

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MDPI and ACS Style

Strasser, L.; Boi, S.; Guapo, F.; Donohue, N.; Barron, N.; Rainbow-Fletcher, A.; Bones, J. Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells. Int. J. Mol. Sci. 2021, 22, 11499. https://doi.org/10.3390/ijms222111499

AMA Style

Strasser L, Boi S, Guapo F, Donohue N, Barron N, Rainbow-Fletcher A, Bones J. Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells. International Journal of Molecular Sciences. 2021; 22(21):11499. https://doi.org/10.3390/ijms222111499

Chicago/Turabian Style

Strasser, Lisa, Stefano Boi, Felipe Guapo, Nicholas Donohue, Niall Barron, Alana Rainbow-Fletcher, and Jonathan Bones. 2021. "Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells" International Journal of Molecular Sciences 22, no. 21: 11499. https://doi.org/10.3390/ijms222111499

APA Style

Strasser, L., Boi, S., Guapo, F., Donohue, N., Barron, N., Rainbow-Fletcher, A., & Bones, J. (2021). Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells. International Journal of Molecular Sciences, 22(21), 11499. https://doi.org/10.3390/ijms222111499

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

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Proteomic Landscape of Adeno-Associated Virus (AAV)-Producing HEK293 Cells

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