Methods of inactivating viral contaminants

Abstract

The present invention relates to a method for preparing an antibody-containing solution free of viral contaminants starting from cultured cells are described. The method include a step of subjecting the antibody containing solution to a mix of solvent and detergent or to high pH.

1 . A method for preparing a virus-inactivated antibody solution, the method comprising: (i) harvesting antibody material produced by transfected cells comprising coding sequences of an antibody, which transfected cells have undergone cell culture, (ii) subjecting said harvested antibody material to a virus inactivation treatment, wherein said virus inactivation treatment is incubation with a mixture of a solvent and a detergent, wherein said harvested antibody material is a clarified harvest. 
     
     
         2 . The method of  claim 1 , wherein said harvested antibody material is produced in non-human mammalian cells. 
     
     
         3 . The method of  claim 1 , wherein said harvested antibody material comprises a monoclonal antibody. 
     
     
         4 . The method of  claim 3 , wherein said monoclonal antibody is a recombinant antibody. 
     
     
         5 . The method of  claim 3 , wherein said monoclonal antibody is multispecific. 
     
     
         6 . The method of  claim 1 , wherein said solvent is TnBP and, wherein said detergent is selected from the group consisting of Triton X-100, Polysorbate 80 and Polysorbate 20. 
     
     
         7 . The method of  claim 6 , wherein the concentration of said TnBP is between about 0.10% (w/w) and about 1% (w/w). 
     
     
         8 . The method of  claim 6 , wherein the concentration of said TnBP is selected from the group consisting of: about 0.1% (w/w), about 0.3% (w/w), about 0.5% (w/w) and about 1% (w/w). 
     
     
         9 . The method of  claim 6 , wherein the concentration of said Polysorbate 80 or said Polysorbate 20 is between about 0.1% (w/w) and about 2% (w/w). 
     
     
         10 . The method of  claim 6 , wherein the concentration of said Polysorbate 80 or said Polysorbate 20 is selected from the group consisting of: about 0.2%, about 0.5% (w/w), about 0.75% (w/w), about 1% (w/w), and about 1.25% (w/w). 
     
     
         11 . The method of  claim 1 , wherein said mixture of a solvent and a detergent is selected from the group consisting of a mixture of 0.3% (w/w) TnBP and 0.5% (w/w) Polysorbate 80, and a mixture of 0.3% (w/w) TnBP and 1% (w/w) Polysorbate 80. 
     
     
         12 . The method of  claim 11 , wherein said harvested antibody material is incubated with a mixture of 0.3% (w/w) TnBP and 1% (w/w) Polysorbate 80 for at least 5 minutes. 
     
     
         13 . The method of  claim 11 , wherein said harvested antibody material is first subjected to protein A chromatography, and wherein a resulting protein A eluate is incubated with a mixture of 0.3% (w/w) TnBP and 1% (w/w) Polysorbate 80 for at least 10 minutes at room temperature. 
     
     
         14 . The method of  claim 11 , wherein said harvested antibody material is incubated with a mixture of 0.3% (w/w) TnBP and 1% (w/w) Polysorbate 80 for about 60 minutes, under agitation, at room temperature. 
     
     
         15 - 20 . (canceled) 
     
     
         21 . The method of  claim 1 , comprising a further step (iii) of testing a portion of the virus-inactivated antibody solution with a viral inactivation assay. 
     
     
         22 . A method of production of a bulk drug substance, the method comprising:
 (a) culturing host cells which have been transfected with coding sequences of an antibody so as to express the antibody;   (b) harvesting a cell culture containing the antibody to obtain a harvested antibody material;   (c) subjecting of the harvested antibody material to viral inactivation by incubating with a mixture of solvent and detergent, wherein said harvested antibody material is a clarified harvest material and is incubated with a mixture of 0.3% (w/w) TnBP and 1% (w/w) Polysorbate 80 for about 60 minutes, under agitation, at room temperature;   (d) subjecting the resulting viral inactivated solution to protein A chromatography;   (e) neutralizing the protein A eluate to pH 6.0, followed by passing the neutralized protein A eluate through a 0.2 μm filter;   (f) subjecting the neutralized protein A filtrate to cation exchange chromatography to produce a cation exchange chromatography eluate, followed by passing the cation exchange chromatography eluate through a 0.2 μm filter;   (g) concentrating the cation exchange chromatography filtrate by ultrafiltration and continuous diafiltration, followed by passing the concentrated ultra-diafiltrate through a 0.2 μm filter;   (h) purifying the concentrated ultra-diafiltrate by anion exchange chromatography in flow through mode, using membrane adsorption, followed by passing the anion exchange chromatography eluate through a 0.2 μm filter;   (i) subjecting the purified anion exchange chromatography filtrate to virus nanofiltration;   (j) concentrating the nanofiltrate by ultrafiltration and continuous diafiltration into a pre-formulation buffer, followed by passing the ultra-diafiltrate containing pre-formulation buffer through a 0.2 μm filter;   (k) adding excipient to the filtered, ultra-diafiltrate containing pre-formulation buffer to obtain a bulk drug substance with a concentration of about 6 mg/mL in a final formulation buffer by adding 5 mM Citrate, 15% Sucrose, and 0.06% Polysorbate 80 at pH 5.9, followed by passing the bulk drug substance through a 0.2 μm filter; and   (l) filling the filtered bulk drug substance into sterile bags, followed by freezing and storage at −80±20° C.   
     
     
         23 - 24 . (canceled) 
     
     
         25 . The method of  claim 6 , wherein the concentration of said Polysorbate 80 or said Polysorbate 20 is between about 0.2% (w/w) and about 1.5% (w/w). 
     
     
         26 . The method of  claim 6 , wherein the concentration of said Polysorbate 80 or said Polysorbate 20 is between about 0.75% (w/w) and about 1.25% (w/w). 
     
     
         27 . The method of  claim 6 , wherein the concentration of said Triton X-100 is between about 0.1% (w/w) and about 2% (w/w). 
     
     
         28 . The method of  claim 1 , wherein said mixture of a solvent and a detergent is selected from the group consisting of: a mixture of 0.3% (w/w) TnBP and 0.2% (w/w) Triton X-100, a mixture of 0.3% (w/w) TnBP and 0.5% (w/w) Triton X-100, and a mixture of 0.3% (w/w) TnBP and 1% (w/w) Triton X-100.