Abstract
The purpose of the present study was to examine the efficacy and mechanism of the PAB (para-amino benzamidine) affinity column chromatography, Viresolve NFP virus filtration, pasteurization (60°C heat treatment for 10 h), and lyophilization steps employed in the manufacture of urokinase from human urine as regards the removal and/or inactivation of the hepatitis A virus (HAV). Samples from the relevant stages of the production process were spiked with HAV 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 (TCID50), and the virus reduction factors evaluated. PAB chromatography was found to be an effective step for removing HAV with a log reduction factor of 3.24. HAV infectivity was rarely detected in the urokinase fraction, while most of the HAV infectivity was recovered in the unbound and wash fractions. HAV was completely removed during the vire solve NFP filtration with a log reduction factor of ≥4.60. Pasteurization was also found to be an effective step in inactivating HAV, where the titers were reduced from an initial titer of 7.18 log10 TCID50 to undetectable levels within 10 h of treatment. The log reduction factor achieved during pasteurization was ≥4.76. Lyophilization revealed the lowest efficacy for inactivating HAV with a log reduction factor of 1.48. The cumulative log reduction factor was ≥14.08. Accordingly, these results indicate that the production process for urokinase exhibited a sufficient HAV reducing capacity to achieve a high margin of virus safety.
Similar content being viewed by others
References
Sherry, S. (1987) Thrombolytic therapy in acute myocardial infarction. A perspective.Drugs 33 (Suppl 3): 1–12.
Coulepis, A. G., S. A. Locarnini, N. I. Lehmann, and I. D. Gust (1980) Detection of hepatitis A virus in the feces of patients with naturally acquired infections.J. Infect. Dis. 141: 151–156.
Andrewes, C. (1989)Andrewes' Viruses of Vertebrates, pp. 120–145. Balliere Tindal, London, UK.
Borovec, S., C. Broumis, W. Adcock, R. Fang, and E. Uren (1998) Inactivation kinetics of model and relevant blood-borne viruses by treatment with sodium hydroxide and heat.Biologicals 26: 237–244.
Kim, I. S., Y. W. Choi, S. R. Lee, M. S. Lee, K. H. Huh, and S. Lee (2001) Removal and inactivation of hepatitis A virus during manufacture of a high purity antihemophilic factor VIII concentrate from human plasma.J. Microbiol. 39: 67–73.
Mosley, J. W. and J. Rakela (1999) Foundling viruses and transfusion medicine.Transfusion 39: 1041–1044.
The European Agency for the Evaluation of Medicinal Products: Human Medicines Evaluation Unit. Committee for Proprietary Medicinal Products (CPMP). Note for guidance on virus validation studies: the design, contribution and interpretation of studies validating the inactivation and removal of viruses (CPMP/BWP/268/95).
International Conference on Harmonisation (1998) Guidance on viral safety evaluation of biotechnology products derived from cell lines of human or animal origin; Availability.Federal Resister 63(185): 51074–51084.
Kim, I. S., Y. W. Choi, H. S. Woo, C. E. Chang, and S. Lee (2000) Solvent/detergent inactivation and chromatographic removal of human immunodeficiency virus during the manufacturing of a high purity antihemophilic factor VIII concentrate.J. Microbiol. 38: 187–191.
Kim, I. S., H. G. Eo, C. W. Park, C. E. Chang, and S. Lee (2001) Removal and inactivation of human immunodeficiency virus (HIV-1) by cold ethanol fractionation and pasteurization during the manufacture of albumin and immunoglobulins from human plasma.Biotechnol. Bioprocess Eng. 6: 25–30.
Han, B., J. O. Carlson, S. M. Powers, and S. R. Wickramasinghe (2002) Enhanced virus removal by flocculation and microfiltration.Biotechnol. Bioprocess Eng. 7: 6–9.
Kim, I. S., Y. W. Choi, S. R. Lee, H. B. Cho, H. G. Eo, H. S. Woo, C. E. Chang, and S. Lee (2001) Improvement of virus safety of a human intravenous immunoglobulin by low pH incubation.J. Microbiol. Biotechnol. 11: 619–627.
Arahana, H. (2001) Viral clearance strategies for biopharmaceutical safety: Part 1. General considerations.Bio. Pharm.. February: 32–43.
Kim, I. S., Y. W. Choi, S. R. Lee, H. S. Woo, and S. Lee (2001) Removal and inactivation of viruses during manufacture of a high purity antihemophilic factor VIII concentrate from human plasma.J. Microbiol. Biotechnol. 11: 497–503.
Hamman, J., J. Zou, and B. Horowitz (1994). Removal and inactivation of hepatitis A virus (HAV) during processing of factor VIII concentrates.Vox Sang. 67 (Suppl. 1): 72–77.
Lemon, S. M., P. C. Murphy, A. Smith, J. Zou, J. Hammon, S. Robinson, and B. Horowitz (1994) Removal/ neutralization of hepatitis A virus during manufacture of high purity, solvent/detergent factor VIII concentrate.J. Med. Virol. 43: 44–49.
Adcock, W. L., A. Macgregor, J. R. Davies, M. Hattarki, D. A. Anderson, and N. H. Goss (1998) Chromatographic removal and heat inactivation of hepatitis A virus during manufacture of human albumin.Biotechnol. Appl. Biochem. 28: 85–94.
Arahana, H. (2001) Viral clearance strategies for biopharmaceutical safety. Part 2: Filtration for viral clearance.Bio. Pharm. February: 32–43.
Aranha-Creado, H., J. Peterson, and P. Y. Huang (1998) Clearance of murine leukaemia virus from monoclonal antibody solutions by a hydrophilic PVDF microporous membrane filter.Biologicals 26: 167–172.
Eibl, J., N. Barrett, T. Hämmerle, and F. Dorner (1996) Nanofiltration of immunoglobulin with 35-nm filters fails to remove substantial amounts of HCV.Biologicals 24: 285–287.
Graf, E. G., E. Jander, A. West, H. Pora, and H. Aranha-Creado (1999) Virus removal by filtration.Dev. Biol. Stand. 99: 89–84.
Oshima, K. H., T. T. Evans-strickfaden, and A. K. Highsmith (1998) Comparison of filtration properties of hepatitis B virus, hepatitis C virus and simian, virus 40 using a polyvinylidene fluoride membrane filter.Vox Sang. 75: 181–188.
Oshima, K. H., T. T. Evans-strickfaden, A. K. Highsmith, and E. W. Ades (1998) The use of a microporus polyvinylidene fluoride (PVDF) membrane filter to separate contaminating viral particles from biologically important proteins.Biologicals 24: 137–145.
Roberts, P. (1997) Efficient removal of viruses by a novel polyvinylidene fluoride membrane filter.J. Virol. Methods 65: 27–31.
Troccoli, N., J. Mclver, A. Losikoff, and J. Poiley (1998) Removal of viruses from human intravenous immune globulin by 35 nm nanofiltration.Biologicals 26: 321–329.
Kim, I. S., H. G. Eo, C. E. Chang, and S. Lee (2000) Partitioning and inactivation of viruses by cold ethanol fractionation and pasteurization during manufacture of albumin from human plasma.J. Microbiol. Biotechnol. 10: 858–864.
Heimburger, N. and H. E. Karges (1989) Strategies to produce virus-safe blood derivatives.Curr. Stud. Hematol. Blood Transfus. 56: 23–33.
Nowak, T., M. Niedrig, D. Bernhardt, and J. Hilfenhaus (1993) Inactivation of HIV, HBV, HCV related viruses and other viruses in human plasma derivatives by pasteurization.Dev. Biol. Stand. 81: 169–176.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Kim, I.S., Choi, Y.W., Lee, S.R. et al. Removal and inactivation of hepatitis A virus during manufacture of urokinase from human urine. Biotechnol. Bioprocess Eng. 7, 340–346 (2002). https://doi.org/10.1007/BF02933518
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02933518