Comparison of different bioreactor systems for the production of high titer retroviral vectors

Improved, human-based packaging cell lines allow the production of high-tit er, RCR-free retroviral vectors. The utility of these cell lines for the pr oduction of clinical grade vectors critically depends on the definition of optimal conditions for scaled-up cultures. In this work, a clone derived fr om 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) x 10(6) PFU cm(-3) in small scale cultures and has been evalu ated for growth and vector production in different reactor systems. The per formances of fixed bed reactors [CellCube (Costar) and Celligen (New Brunsw ick)] and stirred tank reactors [microcarriers and clump cultures] were com pared. The cells showed a higher apparent growth rate in the fixed bed reac tor systems than in the suspension systems, probably as a result of the fac t that aggregation and/or formation of clumps led to a reduced viability an d 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 th e fixed bed systems in comparison to the suspension culture systems. The av erage titers obtained ranged from 0.5 to 2.1 x 10(7) PFU cm(-3) for the fix ed bed and microcarrier systems, while the clump cultures produced only(2-5 ) x 10(5) PFU cm(-3). The differences in titers reflect cell densities as w ell 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 g-fold reduct ion of the seeding cell density, led to a 5-fold increased vector productio n rate accompanied by an average titer of 3 x 10(7) PFU cm(-3) (maximum tit er (4-5) x 10(7) PFU cm(-3)) in the fixed bed reactor. The performance eval uation results using mathematical models indicated that the fixed bed biore actor has a higher potential for retroviral vector production because of bo th the higher reactor productivity and the lower sensitivity of productivit y in relation to the changes in final retrovirus titer in the range of 3 x 10(6) to 15 x 10(6) PFU cm(-3).