Viral clearance during the manufacture of urokinase from human urine

Abstract

= vçåÖ=tççå=`Üçá NIO =~åÇ=få=pÉçé=háã P G= )TGGP %TQUU %QTR ;QPIKP -QTGC &GRCTVOGPV QH /QNGEWNCT 5EKGPEG CPF 6GEJPQNQI[ %QNNGIG QH 'PIKPGGTKPI #LQW 7PKXGTUKV[ 5WYQP -QTGC &GRCTVOGPV QH $KQNQIKECN 5EKGPEGU %QNNGIG QH .KHG 5EKGPEG CPF 0CPQVGEJPQNQI[ *CPPCO 7PKXGTUKV[ &CGLGQP -QTGCĜ Äëíê~Åí=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 o C 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 scaledown 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 ≥6.06, ≥4.60, ≥5.44, and ≥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 ≥5.73 for HIV, ≥3.86 for BVDV, ≥6.75 for BHV, and ≥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. © KSBB

Removal and inactivation of viruses during the manufacture of a high-purity antihemophilic factor IX from human plasma

“…These results indicate that the Viresolve NFP filtration process was a robust and effective method for removal of viruses from factor IX preparations. In previous reports, it has also been shown that the Viresolve NFP filtration process is a robust and effective method for removal of viruses from urokinase preparations [32,47].…”

“…All physico-chemical analyses were conducted according to Standard Operating Procedures (SOPs), based on the Korean Pharmacopoeia, European Pharmacopoeia, and USA Pharmacopoeia. [31,32]. C8166 cells were grown in RPMI 1640 medium containing 2% fetal bovine serum (FBS) and LGlutamine.…”

Para-aminobenzamidine linked regenerated cellulose membranes for plasminogen activator purification: Effect of spacer arm length and ligand density

“…Membrane based chromatographic or batch separation processes have been discussed as promising alternatives for some time now [1] as they lead to more robust processes that are also very amenable to inclusion into a manufacturing process [2, 3]. While membranes provide a more easily accessible surface area for protein binding, they also help overcome the limitations of diffusion resistance, high pressure drop, need for high operating pressure/flow rate, and other usual problems related to traditional chromatographic process scale up [4–8].…”