Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells
Abstract Adeno-associated viral (AAV) vector suspensions produced in either human derived HEK cells or in Spodoptera frugiperda (Sf9) insect cells differ in terms of residual host cell components as well as species-specific post-translational modifications displayed on the AAV capsid proteins. Here...
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Nature Portfolio
2023-02-01
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| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-023-28830-7 |
| _version_ | 1811171750758580224 |
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| author | Kirsten Bucher Eduardo Rodríguez-Bocanegra Bernd Wissinger Torsten Strasser Simon J. Clark Andreas L. Birkenfeld Dorothea Siegel-Axel M. Dominik Fischer |
| author_facet | Kirsten Bucher Eduardo Rodríguez-Bocanegra Bernd Wissinger Torsten Strasser Simon J. Clark Andreas L. Birkenfeld Dorothea Siegel-Axel M. Dominik Fischer |
| author_sort | Kirsten Bucher |
| collection | DOAJ |
| description | Abstract Adeno-associated viral (AAV) vector suspensions produced in either human derived HEK cells or in Spodoptera frugiperda (Sf9) insect cells differ in terms of residual host cell components as well as species-specific post-translational modifications displayed on the AAV capsid proteins. Here we analysed the impact of these differences on the immunogenic properties of the vector. We stimulated human plasmacytoid dendritic cells with various lots of HEK cell-produced and Sf9 cell-produced AAV-CMV-eGFP vectors derived from different manufacturers. We found that AAV8-CMV-eGFP as well as AAV2-CMV-eGFP vectors induced lot-specific but not production platform-specific or manufacturer-specific inflammatory cytokine responses. These could be reduced or abolished by blocking toll-like receptor 9 signalling or by enzymatically reducing DNA in the vector lots using DNase. Successful HEK cell transduction by DNase-treated AAV lots and DNA analyses demonstrated that DNase did not affect the integrity of the vector but degraded extra-viral DNA. We conclude that both HEK- and Sf9-cell derived AAV preparations can contain immunogenic extra-viral DNA components which can trigger lot-specific inflammatory immune responses. This suggests that improved strategies to remove extra-viral DNA impurities may be instrumental in reducing the immunogenic properties of AAV vector preparations. |
| first_indexed | 2024-04-10T17:19:14Z |
| format | Article |
| id | doaj.art-41bcc5402a7c4fcaa9356d25c0d466d9 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-04-10T17:19:14Z |
| publishDate | 2023-02-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-41bcc5402a7c4fcaa9356d25c0d466d92023-02-05T12:11:01ZengNature PortfolioScientific Reports2045-23222023-02-0113111310.1038/s41598-023-28830-7Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cellsKirsten Bucher0Eduardo Rodríguez-Bocanegra1Bernd Wissinger2Torsten Strasser3Simon J. Clark4Andreas L. Birkenfeld5Dorothea Siegel-Axel6M. Dominik Fischer7University Eye Hospital, Centre for Ophthalmology, University Hospital TübingenUniversity Eye Hospital, Centre for Ophthalmology, University Hospital TübingenInstitute for Ophthalmic Research, Centre for Ophthalmology, University Hospital Tübingen, University of TübingenUniversity Eye Hospital, Centre for Ophthalmology, University Hospital TübingenUniversity Eye Hospital, Centre for Ophthalmology, University Hospital TübingenInstitute of Diabetes Research and Metabolic Diseases (IDM) of the Helmholtz Center MunichInstitute of Diabetes Research and Metabolic Diseases (IDM) of the Helmholtz Center MunichUniversity Eye Hospital, Centre for Ophthalmology, University Hospital TübingenAbstract Adeno-associated viral (AAV) vector suspensions produced in either human derived HEK cells or in Spodoptera frugiperda (Sf9) insect cells differ in terms of residual host cell components as well as species-specific post-translational modifications displayed on the AAV capsid proteins. Here we analysed the impact of these differences on the immunogenic properties of the vector. We stimulated human plasmacytoid dendritic cells with various lots of HEK cell-produced and Sf9 cell-produced AAV-CMV-eGFP vectors derived from different manufacturers. We found that AAV8-CMV-eGFP as well as AAV2-CMV-eGFP vectors induced lot-specific but not production platform-specific or manufacturer-specific inflammatory cytokine responses. These could be reduced or abolished by blocking toll-like receptor 9 signalling or by enzymatically reducing DNA in the vector lots using DNase. Successful HEK cell transduction by DNase-treated AAV lots and DNA analyses demonstrated that DNase did not affect the integrity of the vector but degraded extra-viral DNA. We conclude that both HEK- and Sf9-cell derived AAV preparations can contain immunogenic extra-viral DNA components which can trigger lot-specific inflammatory immune responses. This suggests that improved strategies to remove extra-viral DNA impurities may be instrumental in reducing the immunogenic properties of AAV vector preparations.https://doi.org/10.1038/s41598-023-28830-7 |
| spellingShingle | Kirsten Bucher Eduardo Rodríguez-Bocanegra Bernd Wissinger Torsten Strasser Simon J. Clark Andreas L. Birkenfeld Dorothea Siegel-Axel M. Dominik Fischer Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells Scientific Reports |
| title | Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells |
| title_full | Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells |
| title_fullStr | Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells |
| title_full_unstemmed | Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells |
| title_short | Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells |
| title_sort | extra viral dna in adeno associated viral vector preparations induces tlr9 dependent innate immune responses in human plasmacytoid dendritic cells |
| url | https://doi.org/10.1038/s41598-023-28830-7 |
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