Viral clearance during the manufacture of urokinase from human urine

• Published in: Biotechnology and bioprocess engineering 2008, 13(1), , pp.25-32
• Main Authors: Choi, Y.W. (Green Cross Corp., Yongin, Republic of Korea), Kim, I.S. (Hannam University, Daejeon, Republic of Korea), E-mail: [e-mail]
• Format: Article
• Language: English
• Subjects: ACTIVADOR DE PLASMINOGENO; ACTIVATEUR DE PLASMINOGENE; Biotechnology; Bovine viral diarrhea virus; Chemistry; Chemistry and Materials Science; Chromatography; Encephalomyocarditis virus; Filtration; Heat treatment; Herpesvirus; HIV; Human immunodeficiency virus; Inactivation; Industrial and Production Engineering; log reduction factor; Manufacturing; Pasteurization; PLASMINOGEN ACTIVATOR; Urine; viral clearance; virus safety; Viruses; 생물공학
• ISSN: 1226-8372; 1976-3816
• DOI: 10.1007/s12257-007-0140-7

The purpose of the present study was to evaluate the efficacies and mechanisms of the PAB (para-amino benzamidine) affinity column chromatography, virus filtration, pasteurization (60℃ heat treatment for 10 h), and lyophilization steps employed in the manufacture of urokinase from human urine with regard to the removal and/or inactivation of human immunodeficiency virus (HIV), bovine viral diarrhoea virus (BVDV), bovine herpes virus (BHV), and murine encephalomyocarditis virus (EMCV). Samples from relevant stages of the production process were spiked with each virus and subjected to scale-down processes mimicking the manufacture of urokinase. Samples were collected at each step, immediately titrated using a 50% tissue culture infectious dose (TCID∧50), and the virus reduction factors evaluated. PAB chromatography was found to be an effective step for removing BVDV, BHV, and EMCV with log reduction factors of 2.79, 6.50, and 5.96, respectively. HIV, BVDV, BHV, and EMCV were completely removed during the Viresolve NFP filtration step with log reduction factors of greater-than or equal to 6.06, greater-than or equal to 4.60, greater-than or equal to 5.44, and greater-than or equal to 6.87, respectively. Pasteurization was also found to be a robust and effective step in inactivating all the viruses tested, since there were no residual viruses detected after the pasteurization process. The log reduction factors achieved by pasteurization were greater-than or equal to 5.73 for HIV, greater-than or equal to 3.86 for BVDV, greater-than or equal to 6.75 for BHV, and greater-than or equal to 5.92 for EMCV. Lyophilization showed significant efficacy for inactivating BVDV, BHV, and EMCV with log reduction factors of 2.69, 1.37, and 4.70, respectively. These results indicate that the production process for urokinase exhibited a sufficient viral reducing capacity to achieve a high margin of virus safety.