Production of poliovirus at high titers for vaccine production

Abstract

Described is a process for producing poliovirus, the process comprising: a) providing a serum-free suspension culture of cells, which are primary human retina (HER) cells that have been immortalized by expression of adenovirus E1 sequences, b) infecting the cells with poliovirus, at a cell density of between 2×10 6 cells/ml and 150×10 6 cells/ml, and c) harvesting poliovirus at a time of between 12 and 48 hours after infection.

1 . A poliovirus bulk useful for preparation of a polio vaccine, the poliovirus bulk comprising culture medium and poliovirus at a titer of at least about 10 9.4  CCID50/ml, and being produced by a process comprising:
 providing a serum-free suspension culture of cells, which are human fetal retina cells that have been immortalized by expression of adenovirus E1 sequences,   infecting the cells with poliovirus, at a cell density of between 2×10 6  cells/ml and 150×10 6  cells/ml, and   harvesting poliovirus at a time of between 12 and 48 hours after infection to obtain the poliovirus preparation having a titer of at least about 10 9.4  CCID 50 /ml.   
     
     
         2 . The poliovirus bulk of  claim 1 , comprising poliovirus at a titer of at least about 10 9.8  CCID50/ml. 
     
     
         3 . The poliovirus bulk of  claim 1 , comprising poliovirus at a titer of at least about 10 10  CCID50/ml. 
     
     
         4 . The poliovirus bulk of  claim 1 , comprising poliovirus type 1, poliovirus type 2, or poliovirus type 3. 
     
     
         5 . The poliovirus bulk of  claim 1 , wherein the poliovirus is a wild-type poliovirus. 
     
     
         6 . The poliovirus bulk of  claim 1 , wherein the poliovirus is an attenuated poliovirus. 
     
     
         7 . The poliovirus bulk of  claim 1 , which has a volume of between 1 and 1000 liters. 
     
     
         8 . The poliovirus bulk of  claim 1 , further comprising cells and/or cell debris of the cells utilized in the process. 
     
     
         9 . The poliovirus bulk of  claim 1 , which is present in a bioreactor 
     
     
         10 . The poliovirus bulk of  claim 1 , which is present in a suitable container. 
     
     
         11 . A process for producing an inactivated poliovirus vaccine, the process comprising:
 purifying and inactivating poliovirus from the poliovirus bulk of  claim 1 .   
     
     
         12 . A polio vaccine free of monkey-derived protein. 
     
     
         13 . The polio vaccine of  claim 12 , which is free of non-human protein. 
     
     
         14 . An inactivated polio vaccine free of non-human protein, produced by a method comprising:
 (i) obtaining a poliovirus preparation in cell culture at a titer of at least about 10 9.4  CCID 50 /ml by a process comprising:
 a) providing a serum-free suspension culture of cells, which are human fetal retina cells that have been immortalized by expression of adenovirus E1 sequences, 
 b) infecting the cells with poliovirus, at a cell density of between 2×10 6  cells/ml and 150×10 6  cells/ml, 
 c) propagating the poliovirus in the cells at a temperature of between 34° C. and 36° C., and 
 d) harvesting poliovirus at a time of between 12 and 48 hours after infection to obtain the poliovirus preparation having a titer of at least about 10 9.4  CCID 50 /ml; 
   (ii) purifying the harvested poliovirus;   (iii) inactivating the purified poliovirus; and   (iv) formulating the inactivated poliovirus to obtain an inactivated poliovirus vaccine.   
     
     
         15 . The inactivated polio vaccine of  claim 14 , wherein the cells utilized in the method are PER.C6® cells. 
     
     
         16 . The inactivated polio vaccine of  claim 14 , wherein the cells in the method are infected at a cell density of between 8×10 6  cells/ml and 50×10 6  cells/ml. 
     
     
         17 . The inactivated polio vaccine of  claim 14 , wherein the cells in the method are cultured at a temperature of between 34° C. and 36° C. 
     
     
         18 . The inactivated polio vaccine of  claim 14 , comprising poliovirus type 1, poliovirus type 2, and poliovirus type 3. 
     
     
         19 . The inactivated polio vaccine of  claim 14 , wherein the poliovirus is a wild-type poliovirus. 
     
     
         20 . The inactivated polio vaccine of  claim 14 , wherein the poliovirus is an attenuated poliovirus.