A replication-competent adenovirus assay for E1-deleted Ad35 vectors produced in PER.C6 cells

G. Marzio; E. Kerkvliet; J. A. Bogaards; S. Koelewijn; A. De Groot; L. Gijsbers; G. J. Weverling; R. Vogels; M. Havenga; J. Custers; M. G. Pau; J. Y. Guichoux; J. Lewis; J. Goudsmit

doi:https://doi.org/10.1016/j.vaccine.2006.12.011

A replication-competent adenovirus assay for E1-deleted Ad35 vectors produced in PER.C6 cells

G. Marzio, E. Kerkvliet, J. A. Bogaards, S. Koelewijn, A. De Groot, L. Gijsbers, G. J. Weverling, R. Vogels, M. Havenga, J. Custers, M. G. Pau, J. Y. Guichoux, J. Lewis, J. Goudsmit

Research output: Contribution to journal › Article › Academic › peer-review

12 Citations (Scopus)

Abstract

The presence of replication-competent adenovirus (RCA) is a safety concern for biologics based on recombinant adenoviruses and RCA testing is therefore mandatory for release of clinical material. RCA, which arises from homologous recombination between Ad5 vectors and HEK-293 cells, can be eliminated by the use of PER.C6 cells in combination with a matched vector. However, little is known on RCA formation with vectors based on adenovirus serotypes other than Ad5 and reliable RCA assays to test them are generally lacking. Here we report on the development and qualification of a sensitive RCA assay for Ad35, a promising alternative to Ad5 vectors. The assay is able to detect 1 RCA in 3 × 10¹⁰ vector particles with 95% confidence, thus meeting current FDA requirements, and can discriminate between RCA and other rare CPE-causing entities, including helper dependent E1 positive particles (HDEP). Using this assay, the first batches of Ad35 vectors produced in PER.C6 cells were analysed and found to be free of RCA and HDEP. Based on the statistical model used, we anticipate that our approach to RCA assay development can be broadly applicable to other adenoviral vectors.

Original language	English
Pages (from-to)	2228-2237
Number of pages	10
Journal	Vaccine
Volume	25
Issue number	12
DOIs	https://doi.org/10.1016/j.vaccine.2006.12.011
Publication status	Published - 8 Mar 2007

Keywords

Adenovirus
Assay
HDEP
PER.C6
RCA

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Marzio, G., Kerkvliet, E., Bogaards, J. A., Koelewijn, S., De Groot, A., Gijsbers, L., Weverling, G. J., Vogels, R., Havenga, M., Custers, J., Pau, M. G., Guichoux, J. Y., Lewis, J., & Goudsmit, J. (2007). A replication-competent adenovirus assay for E1-deleted Ad35 vectors produced in PER.C6 cells. Vaccine, 25(12), 2228-2237. https://doi.org/10.1016/j.vaccine.2006.12.011

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title = "A replication-competent adenovirus assay for E1-deleted Ad35 vectors produced in PER.C6 cells",

abstract = "The presence of replication-competent adenovirus (RCA) is a safety concern for biologics based on recombinant adenoviruses and RCA testing is therefore mandatory for release of clinical material. RCA, which arises from homologous recombination between Ad5 vectors and HEK-293 cells, can be eliminated by the use of PER.C6 cells in combination with a matched vector. However, little is known on RCA formation with vectors based on adenovirus serotypes other than Ad5 and reliable RCA assays to test them are generally lacking. Here we report on the development and qualification of a sensitive RCA assay for Ad35, a promising alternative to Ad5 vectors. The assay is able to detect 1 RCA in 3 × 1010 vector particles with 95% confidence, thus meeting current FDA requirements, and can discriminate between RCA and other rare CPE-causing entities, including helper dependent E1 positive particles (HDEP). Using this assay, the first batches of Ad35 vectors produced in PER.C6 cells were analysed and found to be free of RCA and HDEP. Based on the statistical model used, we anticipate that our approach to RCA assay development can be broadly applicable to other adenoviral vectors.",

keywords = "Adenovirus, Assay, HDEP, PER.C6, RCA",

author = "G. Marzio and E. Kerkvliet and Bogaards, {J. A.} and S. Koelewijn and {De Groot}, A. and L. Gijsbers and Weverling, {G. J.} and R. Vogels and M. Havenga and J. Custers and Pau, {M. G.} and Guichoux, {J. Y.} and J. Lewis and J. Goudsmit",

note = "Funding Information: This Project has been funded in part with Federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, under Contract No. AI-05421 for Science Applications International Corporation.",

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doi = "https://doi.org/10.1016/j.vaccine.2006.12.011",

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T1 - A replication-competent adenovirus assay for E1-deleted Ad35 vectors produced in PER.C6 cells

AU - Marzio, G.

AU - Kerkvliet, E.

AU - Bogaards, J. A.

AU - Koelewijn, S.

AU - De Groot, A.

AU - Gijsbers, L.

AU - Weverling, G. J.

AU - Vogels, R.

AU - Havenga, M.

AU - Custers, J.

AU - Pau, M. G.

AU - Guichoux, J. Y.

AU - Lewis, J.

AU - Goudsmit, J.

N1 - Funding Information: This Project has been funded in part with Federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, under Contract No. AI-05421 for Science Applications International Corporation.

PY - 2007/3/8

Y1 - 2007/3/8

N2 - The presence of replication-competent adenovirus (RCA) is a safety concern for biologics based on recombinant adenoviruses and RCA testing is therefore mandatory for release of clinical material. RCA, which arises from homologous recombination between Ad5 vectors and HEK-293 cells, can be eliminated by the use of PER.C6 cells in combination with a matched vector. However, little is known on RCA formation with vectors based on adenovirus serotypes other than Ad5 and reliable RCA assays to test them are generally lacking. Here we report on the development and qualification of a sensitive RCA assay for Ad35, a promising alternative to Ad5 vectors. The assay is able to detect 1 RCA in 3 × 1010 vector particles with 95% confidence, thus meeting current FDA requirements, and can discriminate between RCA and other rare CPE-causing entities, including helper dependent E1 positive particles (HDEP). Using this assay, the first batches of Ad35 vectors produced in PER.C6 cells were analysed and found to be free of RCA and HDEP. Based on the statistical model used, we anticipate that our approach to RCA assay development can be broadly applicable to other adenoviral vectors.

AB - The presence of replication-competent adenovirus (RCA) is a safety concern for biologics based on recombinant adenoviruses and RCA testing is therefore mandatory for release of clinical material. RCA, which arises from homologous recombination between Ad5 vectors and HEK-293 cells, can be eliminated by the use of PER.C6 cells in combination with a matched vector. However, little is known on RCA formation with vectors based on adenovirus serotypes other than Ad5 and reliable RCA assays to test them are generally lacking. Here we report on the development and qualification of a sensitive RCA assay for Ad35, a promising alternative to Ad5 vectors. The assay is able to detect 1 RCA in 3 × 1010 vector particles with 95% confidence, thus meeting current FDA requirements, and can discriminate between RCA and other rare CPE-causing entities, including helper dependent E1 positive particles (HDEP). Using this assay, the first batches of Ad35 vectors produced in PER.C6 cells were analysed and found to be free of RCA and HDEP. Based on the statistical model used, we anticipate that our approach to RCA assay development can be broadly applicable to other adenoviral vectors.

KW - Adenovirus

KW - Assay

KW - HDEP

KW - PER.C6

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ER -

A replication-competent adenovirus assay for E1-deleted Ad35 vectors produced in PER.C6 cells

Abstract

Keywords

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