Method for producing recombinant adenovirus

Abstract

The invention concerns a method for producing recombinant adenovirus by which viral DNA is introduced in a packaging cell culture and the viruses produced are harvested after liberation in the supernatant. The invention also concerns the viruses produced and their use.

1 - 32 . (canceled)  
     
     
         33 . A purified adenovirus composition, wherein the composition is essentially free of contaminating proteins as determined by its absorbance ratio A 260 nm /A 280 nm , western blot, or HPLC.  
     
     
         34 . The purified adenovirus composition of  claim 33 , wherein the composition has an absorbance ratio A 260 nm /A 280 nm  equal to 1.30+/−0.05.  
     
     
         35 . The purified adenovirus composition of  claim 33 , wherein the composition is essentially free of contaminating bovine serum albumin (BSA).  
     
     
         36 . The purified adenovirus composition of  claim 35 , wherein the contaminating BSA content in the composition is less than 100 ng per mg of virus as determined by western blot analysis with an anti-BSA polyclonal antibody.  
     
     
         37 . The purified adenovirus composition of  claim 35 , wherein the contaminating BSA content in the composition is less than 0.1% as determined by HPLC.  
     
     
         38 . A purified adenovirus composition, wherein the composition is essentially free of contaminating nucleic acids as determined by its absorbance ratio A 260 nm /A 280 nm  or by electrophoretic analysis.  
     
     
         39 . The purified adenovirus composition of  claim 38 , wherein the composition has an absorbance ratio A 260 nm /A 280 nm  equal to 1.30+/−0.05.  
     
     
         40 . The purified adenovirus composition of  claim 38 , wherein the composition is essentially free of contaminating nucleic acid as determined by electorphoretic analysis.  
     
     
         41 . A purified adenovirus composition, wherein the composition forms a single band of density 1.30 upon CsCl gradient ultracentrifugation.  
     
     
         42 . The purified adenovirus composition of  claim 41 , wherein the composition does not comprise empty virus particles.  
     
     
         43 . The purified adenovirus composition of  claim 41 , wherein the composition does not comprise capsid fragments.  
     
     
         44 . The purified adenovirus composition of  claim 41 , wherein the composition does not comprise aggregated virus particles.  
     
     
         45 . A purified adenovirus composition, wherein the composition is purified 70 fold in terms of quantity of total proteins when compared to the same composition before a one-step ion-exchange purification.  
     
     
         46 . A purified adenovirus composition, wherein the composition has a ratio of number of viral particles measured by HPLC to the number of plaque-forming units (pfu) that is between 16 and 78.  
     
     
         47 . A purified recombinant adenovirus composition, wherein the adenovirus composition is produced by a process comprising: 
 (a) introducing adenoviral DNA into a culture of encapsidation cells; and    (b) harvesting adenoviruses following their release into the culture supernatant without lysis of the encapsidation cells by an external factor.    
     
     
         48 . A purified adenovirus composition, wherein the adenovirus composition is produced by a process comprising: 
 (a) obtaining a biological medium comprising adenovirus;    (b) performing a single chromatography step comprising anion-exchange chromatography on the biological medium; and    (c) recovering the adenovirus with an adenoviral particle yield of at least 70% of the viral particles present in the biological medium prior to chromatography.