Herpes Simplex Virus Clearance During Purification of a Recombinant Adeno-Associated Virus Serotype 1 Vector
Abstract
Gene delivery vectors based on adeno-associated virus (AAV) have potential utility for treatment of many genetic disorders. Current AAV vector manufacturing methods employ helper viruses to deliver functions needed to produce replication-defective recombinant AAV (rAAV) vectors, and clearance of infectious helper virus from the drug substance is essential for ensuring the safety of rAAV-based therapies. We have developed a manufacturing method for the production of rAAV vectors using a recombinant herpes simplex virus type 1 (rHSV) complementation system in suspension baby hamster kidney cells. The manufacturing process includes three primary unit operations, detergent lysis of the cell harvest and two downstream column chromatography steps, which achieve viral clearance. These unit operations inactivate and remove HSV, including replication-competent HSV present at low levels in rHSV helper stocks. Here we report results quantifying the reduction in HSV achieved during rAAV vector purification. Clearance of HSV was at least 6.84 log10 with 1% Triton X-100, 4.34 log10 with CIM Q column chromatography, and 2.86 log10 with AVB affinity chromatography. Combined, these three orthogonal methods achieved clearance of at least 14.04 log10 of HSV. The total input quantity of rHSV in a 100-liter production batch is approximately 1.2×10(12) plaque-forming units (pfu), and after purification, the concentration of residual rHSV in the resulting drug substance of approximately 450 ml woul…
Integrity signals
Registry
Cites
Statistics
Claims the paper makes
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
Approaches to develop successful virus tests have been developed and used.
“While challenging to test for adventitious viruses in the presence of recombinant viral stocks, approaches to develop successful virus tests have been developed and used 53 .”
From Viral contamination in biologic manufacture and implications for emerging therapies (2020) · cites this paper as [53]
The IVV assay takes between 14 to 28 days to complete.
“An additional challenge is that the IVV assay takes between 14 to 28 days to complete 38,53,59,60 and is not amenable to the rapid release required of some ATMP products.”
From Viral contamination in biologic manufacture and implications for emerging therapies (2020) · cites this paper as [53]
Cited by
Sorted by citations
Manufacturing of recombinant adeno-associated viral vectors for clinical trials
Clément, N., & Grieger, J. C. (2016) · Molecular Therapy - Methods & Clinical Development · doi:10.1038/mtm.2016.2 · Cited by 286 · see citing works
Viral contamination in biologic manufacture and implications for emerging therapies
Barone, P. W., Wiebe, M. E., Leung, J. C., Hussein, I. T. M., Keumurian, F. J., Bouressa, J., Brussel, A., Chen, D., Chong, M., Dehghani, H., et al. (2020) · Nature Biotechnology · doi:10.1038/s41587-020-0507-2 · Cited by 212 · see citing works
Pharmacology of Recombinant Adeno-associated Virus Production
Penaud-Budloo, M., François, A., Clément, N., & Ayuso, E. (2018) · Molecular Therapy - Methods & Clinical Development · doi:10.1016/j.omtm.2018.01.002 · Cited by 177 · see citing works
AAV Production Using Baculovirus Expression Vector System
Sandro, Q., Relizani, K., & Benchaouir, R. (2019) · Methods in Molecular Biology · doi:10.1007/978-1-4939-9065-8_5 · Cited by 24 · see citing works
- Authors
- Ye, G.-J., Scotti, M. M., Thomas, D. L., Wang, L., Knop, D. R., & Chulay, J. D.
- Affiliations
- Applied Genetic Technologies (United States) (US)
- ORCID
- Wang, Lijun
- Venue
- Human Gene Therapy Clinical Development
- Publisher
- SAGE Publications
- Published
- 2014-12
- Volume
- 25
- Issue
- 4
- Pages
- 212-217
- ISSN
- 2324-8637 (ISSN-L)
- 2324-8645
- OpenAlex
- W2026979258
- Cited by
- 21
- References
- 0
- Citations by year
- 2025: 1 · 2024: 1 · 2023: 5 · 2022: 1 · 2021: 1 · 2020: 2 · 2019: 2 · 2018: 2 · 2017: 3 · 2016: 2 · 2015: 1
- FWCI
- 1.31
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Human Gene Therapy Clinical Development (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Viral Infectious Diseases and Gene Expression in Insects
- Keywords
- recombinant adeno-associated virus, herpes simplex virus type 1, viral clearance, detergent lysis, Triton X-100, affinity chromatography, column chromatography, BHK-21 cell culture
- MeSH terms
- Dependovirus, Animals, Cell Line, Chlorocebus aethiops, DNA, Recombinant, Genetic Engineering, Genetic Vectors, Cricetinae, Vero Cells, Herpesvirus 1, Human
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
