Incubation Temperature and Period During Denarase Treatment and Microfiltration Affect the Yield of Recombinant Adenoviral Vectors During Downstream Processing

Sonugür F. G., Babahan C., Abdi Abgarmi S., Akbulut H.

MOLECULAR BIOTECHNOLOGY, cilt.65, sa.7, ss.1129-1139, 2023 (SCI-Expanded) identifier identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 65 Sayı: 7
  • Basım Tarihi: 2023
  • Doi Numarası: 10.1007/s12033-022-00616-8
  • Dergi Adı: MOLECULAR BIOTECHNOLOGY
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, BIOSIS, Biotechnology Research Abstracts, CAB Abstracts, Chemical Abstracts Core, Compendex, Food Science & Technology Abstracts, Index Islamicus, INSPEC, MEDLINE, Veterinary Science Database
  • Sayfa Sayıları: ss.1129-1139
  • Anahtar Kelimeler: Downstream processing, Bioprocess development, Viruses, Vaccines, SARS-CoV-2, VIRAL VECTORS, TRITON X-100, PURIFICATION, STABILITY, PRODUCT, CLARIFICATION, REPLICATION, FORMULATION, CHALLENGES, FILTRATION
  • Ankara Üniversitesi Adresli: Evet

Özet

Adenoviral vectors (AV) are commonly used as vaccine and gene therapy vehicles because of their ease of construction, ability to grow to high titers in the large-scale production process, and safety for human applications. However, the efficiency rate of downstream processes for adenoviral vectors still varies greatly. In the current study, we aimed to investigate the effect of the downstream treatment protocol and microfiltration of the harvested upstream material on viral vector yield. We compared the performance of the repeated freeze-thaw (RFT) and the Tween-20 detergent lysis (DLT) methods. In addition, the effects of the cell lysis method, incubation temperature, and time on viral yield were investigated. The samples were incubated at either room temperature or 37 & DEG;C for 1-, 2-, and 4-h periods. Samples were filtered with PES and SFCA membrane. Virus yield and infectivity were assayed by qPCR and immuno-titration. In conclusion, our results suggest that 2-h incubation gives the best results when incubated at 37 & DEG;C for denarase activity when Tween-20 is used for virus recovery. If the room temperature is preferred, 4-h incubation could be preferred. A phase 1 clinical trial (NCT05526183, January 21, 2022) was started with the recombinant adenovirus used in the study.