KINETICS OF RETROVIRAL PRODUCTION FROM THE AMPHOTROPIC PSI-CRIP MURINE PRODUCER CELL-LINE

Rapidly expanding development and practice of gene therapy requires th e availability of large quantities of high titer retroviral supernatan ts. One way to achieve high retroviral titers is through improved unde rstanding of the kinetics of retroviral production and decay, and the subsequent development of improved cell culture methods. In the presen t study we investigated the effects of different operational modes on the retroviral production of the NIH 3T3 fibroblast derived amphotropi c murine retroviral producing cell line pMFG/Psi CRIP. Semi-continuous culture (exchange of 50% of medium volume daily) was found to promote cell growth and enhance retroviral production. The rapid medium excha nge resulted in significantly larger amounts of high titer supernatant s and an extended production phase as compared to the batch control cu ltures. The specific viral productivity of the pMFG/Psi CRIP cells was in the range of 10 to 40 infectious viruses produced per thousand pro ducer cells per day. The CV-1 African Green Monkey kidney cell line wa s used as the infection target. Lowering the serum level from 20% to 1 0% improved retroviral production slightly. However, at lower serum le vels (1%, 5% and 10% (v/v)) growth of the producer cell line, and thus retroviral production, was directly proportional to the serum level. The half-life of the virus at 37 degrees C was found to be 5.5 hours. Promoting the growth of producer cell lines can improve retroviral vec tors titers and viral production. High cell density systems that allow for rapid cell growth and waste product removal are likely to be used to generate high-titer retroviral supernatants.