Engineered CHO cells as a novel AAV production platform for gene therapy delivery
Abstract The Herpes simplex virus (HSV)-based platform for production of recombinant adeno-associated viral vectors (rAAVs) yields higher titers and increased percentage of full capsids when compared to the triple transient transfection (TTT) method. However, this platform currently faces two major...
| Main Authors: | , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Nature Portfolio
2023-11-01
|
| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-023-46298-3 |
| _version_ | 1797630211083206656 |
|---|---|
| author | Abdou Nagy Lina Chakrabarti James Kurasawa Sri Hari Raju Mulagapati Paul Devine Jamy Therres Zhongying Chen Albert E. Schmelzer |
| author_facet | Abdou Nagy Lina Chakrabarti James Kurasawa Sri Hari Raju Mulagapati Paul Devine Jamy Therres Zhongying Chen Albert E. Schmelzer |
| author_sort | Abdou Nagy |
| collection | DOAJ |
| description | Abstract The Herpes simplex virus (HSV)-based platform for production of recombinant adeno-associated viral vectors (rAAVs) yields higher titers and increased percentage of full capsids when compared to the triple transient transfection (TTT) method. However, this platform currently faces two major challenges. The first challenge is the reliance on commercial media, sometimes supplemented with serum, leading to costly manufacturing and a high risk for introduction of adventitious agents. The second challenge is that the production of HSV-1 relies on adherent complementing Vero cells (V27), making it difficult to scale up. We engineered serum-free-adapted CHO cells expressing key HSV-1 entry receptors, HVEM and/or Nectin-1 to address the first challenge. Using high-throughput cloning methods, we successfully selected a HVEM receptor-expressing clone (CHO–HV–C1) that yields 1.62 × 109, 2.51 × 109, and 4.07 × 109 viral genome copies/mL with rAAV6.2-GFP, rAAV8-GFP, and rAAV9-GFP vectors respectively, within 24 h post rHSV-1 co-infection. Moreover, CHO–HV–C1-derived rAAVs had comparable in vitro transduction, infectivity, and biodistribution titers to those produced by TTT. The second challenge was addressed via engineering CHO–HV–C1 cells to express HSV-1 CP27. These cells successfully produced rHSV-1 vectors, but with significantly lower titers than V27 cells. Taken together, the CHO/HSV system provides a novel, scalable, reduced cost, serum-free AAV manufacturing platform. |
| first_indexed | 2024-03-11T11:04:01Z |
| format | Article |
| id | doaj.art-05815a7b40d14da2996bba9473daed83 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-03-11T11:04:01Z |
| publishDate | 2023-11-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-05815a7b40d14da2996bba9473daed832023-11-12T12:17:02ZengNature PortfolioScientific Reports2045-23222023-11-0113111610.1038/s41598-023-46298-3Engineered CHO cells as a novel AAV production platform for gene therapy deliveryAbdou Nagy0Lina Chakrabarti1James Kurasawa2Sri Hari Raju Mulagapati3Paul Devine4Jamy Therres5Zhongying Chen6Albert E. Schmelzer7Cell Culture and Fermentation Sciences, Biopharmaceutical Development, BioPharmaceuticals R&D, AstraZenecaCell Culture and Fermentation Sciences, Biopharmaceutical Development, BioPharmaceuticals R&D, AstraZenecaBiologics Engineering, R&D, AstraZenecaAnalytical Science, Biopharmaceutical Development, Biopharma R&D, AstraZenecaAnalytical Science, Biopharmaceutical Development, Biopharma R&D, AstraZenecaCell Culture and Fermentation Sciences, Biopharmaceutical Development, BioPharmaceuticals R&D, AstraZenecaClinical Pharmacology and Safety Sciences, AstraZenecaCell Culture and Fermentation Sciences, Biopharmaceutical Development, BioPharmaceuticals R&D, AstraZenecaAbstract The Herpes simplex virus (HSV)-based platform for production of recombinant adeno-associated viral vectors (rAAVs) yields higher titers and increased percentage of full capsids when compared to the triple transient transfection (TTT) method. However, this platform currently faces two major challenges. The first challenge is the reliance on commercial media, sometimes supplemented with serum, leading to costly manufacturing and a high risk for introduction of adventitious agents. The second challenge is that the production of HSV-1 relies on adherent complementing Vero cells (V27), making it difficult to scale up. We engineered serum-free-adapted CHO cells expressing key HSV-1 entry receptors, HVEM and/or Nectin-1 to address the first challenge. Using high-throughput cloning methods, we successfully selected a HVEM receptor-expressing clone (CHO–HV–C1) that yields 1.62 × 109, 2.51 × 109, and 4.07 × 109 viral genome copies/mL with rAAV6.2-GFP, rAAV8-GFP, and rAAV9-GFP vectors respectively, within 24 h post rHSV-1 co-infection. Moreover, CHO–HV–C1-derived rAAVs had comparable in vitro transduction, infectivity, and biodistribution titers to those produced by TTT. The second challenge was addressed via engineering CHO–HV–C1 cells to express HSV-1 CP27. These cells successfully produced rHSV-1 vectors, but with significantly lower titers than V27 cells. Taken together, the CHO/HSV system provides a novel, scalable, reduced cost, serum-free AAV manufacturing platform.https://doi.org/10.1038/s41598-023-46298-3 |
| spellingShingle | Abdou Nagy Lina Chakrabarti James Kurasawa Sri Hari Raju Mulagapati Paul Devine Jamy Therres Zhongying Chen Albert E. Schmelzer Engineered CHO cells as a novel AAV production platform for gene therapy delivery Scientific Reports |
| title | Engineered CHO cells as a novel AAV production platform for gene therapy delivery |
| title_full | Engineered CHO cells as a novel AAV production platform for gene therapy delivery |
| title_fullStr | Engineered CHO cells as a novel AAV production platform for gene therapy delivery |
| title_full_unstemmed | Engineered CHO cells as a novel AAV production platform for gene therapy delivery |
| title_short | Engineered CHO cells as a novel AAV production platform for gene therapy delivery |
| title_sort | engineered cho cells as a novel aav production platform for gene therapy delivery |
| url | https://doi.org/10.1038/s41598-023-46298-3 |
| work_keys_str_mv | AT abdounagy engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT linachakrabarti engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT jameskurasawa engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT sriharirajumulagapati engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT pauldevine engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT jamytherres engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT zhongyingchen engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery AT alberteschmelzer engineeredchocellsasanovelaavproductionplatformforgenetherapydelivery |