Comparison of Different Bioreactor Systems for the Production of High Titer Retroviral Vectors

• Published in: Biotechnology progress Vol. 17; no. 2; pp. 326 - 335
• Main Authors: Merten, O.-W., Cruz, P. E., Rochette, C., Geny-Fiamma, C., Bouquet, C., Gonçalves, D., Danos, O., Carrondo, M. J. T.
• Format: Journal Article
• Language: English
• Published: USA American Chemical Society 01-03-2001; American Institute of Chemical Engineers
• Subjects: Animal cells; Biological and medical sciences; Bioreactors; Biotechnology; Cell Line; Eukaryotic cell cultures; Fundamental and applied biological sciences. Psychology; Genetic Vectors - biosynthesis; gibbon ape leukemia virus; Methods. Procedures. Technologies; Miscellaneous; Retroviridae - genetics; Retrovirus; Various methods and equipments; Performance evaluation; Human; Productivity; Cell culture; Packed bed; Fixed bed reactor; Retroviridae; Virus; Bioreactor; Cell line; Animal; Production; Mathematical model; Vector; Comparative study
• ISSN: 8756-7938; 1520-6033
• DOI: 10.1021/bp000162z

Improved, human‐based packaging cell lines allow the production of high‐titer, RCR‐free retroviral vectors. The utility of these cell lines for the production of clinical grade vectors critically depends on the definition of optimal conditions for scaled‐up cultures. In this work, a clone derived from the TE Fly GALV packaging cell (Duisit et al. Hum. Gene Ther. 1999, 10, 189) that produces high titers of a lacZ containing retroviral vector with a Gibbon Ape Leukemia Virus envelope glycoprotein was used. This clone can produce (2−5) × 106 PFU cm−3 in small scale cultures and has been evaluated for growth and vector production in different reactor systems. The performances of fixed bed reactors [CellCube (Costar) and Celligen (New Brunswick)] and stirred tank reactors [microcarriers and clump cultures] were compared. The cells showed a higher apparent growth rate in the fixed bed reactor systems than in the suspension systems, probably as a result of the fact that aggregation and/or formation of clumps led to a reduced viability and reduced growth of cells in the interior of the clumps. As a consequence, the final cell density and number were in average 3‐ to 7‐fold higher in the fixed bed systems in comparison to the suspension culture systems. The average titers obtained ranged from 0.5 to 2.1 × 107 PFU cm−3 for the fixed bed and microcarrier systems, while the clump cultures produced only (2−5) × 105 PFU cm−3. The differences in titers reflect cell densities as well as specific viral vector production rates, with the immobilization and microcarrier systems exhibiting an at least 10‐fold higher production rate in comparison to the clump cultures. A partial optimization of the culture conditions in the Celligen fixed bed reactor, consisting of a 9‐fold reduction of the seeding cell density, led to a 5‐fold increased vector production rate accompanied by an average titer of 3 × 107 PFU cm−3 (maximum titer (4−5) × 107 PFU cm−3) in the fixed bed reactor. The performance evaluation results using mathematical models indicated that the fixed bed bioreactor has a higher potential for retroviral vector production because of both the higher reactor productivity and the lower sensitivity of productivity in relation to the changes in final retrovirus titer in the range of 3 × 106 to 15 × 106 PFU cm−3.