Development of a measles vaccine production process in MRC-5 cells grown on Cytodex1 microcarriers and in a stirred bioreactor

摘要

Measles vaccination remains the most efficient way to control the spread of the virus. This work focuses on the production of a measles vaccine using stirred conditions as an advanced option for process scale up. Non-porous Cytodex 1 microcarriers were used to support MRC-5 cell growth in suspension cultures. Virus replication was first optimized in spinner flasks, and the effects of various operational parameters were investigated. Cell infection with AIK-C measles strain at an MOI (multiplicity of infection) of 0.005, without glucose regulation and in M199 medium, resulted in a virus titer of 10 ~(6.25) TCID _(50) (median tissue culture infective dose)/ml. To optimize the production process in a 7-l bioreactor, we carried out various perfused cultures using minimum essential medium (MEM)+5% FCS diluted with phosphate-buffered saline (PBS). We achieved a high cell density level (4.1∈×∈10 6 cells/ml) with an efficient use of the medium when MEM+5% FCS diluted with PBS at 25% was used during the cell amplification step. Optimization of measles production in MRC-5 cells grown on Cytodex 1 beads in a 7-l bioreactor showed that perfusion was the most efficient when compared to repeated-batch culture. Perfusion at a rate of 0.25 V (reactor volume)/day showed the highest specific productivity (1.6 IVP [infectious virus particle] cell ~(-1) day ~(-1)). Testing of several stabilizers containing pharmaceutically improved components such as sugars, amino acids, and charged ions showed that the formulation composed of sucrose and MgCl _2, led to the maintenance of the infectivity of the AIK-C measles virus strain to a significant level, when stored at +28°C, +4°C and -60°C.