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Partitioning of human and sheep forms of the pathogenic prion protein during the purification of therapeutic proteins from human plasma

Authors

  • Christopher J. Stenland,

    1. From the Department of Pathogen Safety Research, Bayer Biological Products, Research Triangle Park, North Carolina
    2. Laboratory of CNS Studies, National Institutes of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland
    3. Laboratory of Molecular and Biochemical Neurovirology, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York.
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  • Douglas C. Lee,

    1. From the Department of Pathogen Safety Research, Bayer Biological Products, Research Triangle Park, North Carolina
    2. Laboratory of CNS Studies, National Institutes of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland
    3. Laboratory of Molecular and Biochemical Neurovirology, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York.
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  • Paul Brown,

    1. From the Department of Pathogen Safety Research, Bayer Biological Products, Research Triangle Park, North Carolina
    2. Laboratory of CNS Studies, National Institutes of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland
    3. Laboratory of Molecular and Biochemical Neurovirology, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York.
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  • Stephen R. Petteway Jr.,

    1. From the Department of Pathogen Safety Research, Bayer Biological Products, Research Triangle Park, North Carolina
    2. Laboratory of CNS Studies, National Institutes of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland
    3. Laboratory of Molecular and Biochemical Neurovirology, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York.
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  • Richard Rubenstein

    1. From the Department of Pathogen Safety Research, Bayer Biological Products, Research Triangle Park, North Carolina
    2. Laboratory of CNS Studies, National Institutes of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland
    3. Laboratory of Molecular and Biochemical Neurovirology, New York State Institute for Basic Research in Developmental Disabilities, Staten Island, New York.
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  • ABBREVIATIONS:

    GSS = Gerstmann-Sträussler-Scheinker syndrome; PK = proteinase K; PrP(s) = prion protein(s); RES = proteinase resistant; Sc = scrapie; sCJD = sporadic CJD; TSE = transmissible spongiform encephalopathy; vCJD = variant CJD.

  • This work was supported in part by the Office of Mental Retardation in Developmental Disabilities.

Address reprint requests to: Chris Stenland, PhD, Department of Pathogen Safety Research, Bayer Corporation, PO Box 13887, 85 T. W. Alexander Drive, Research Triangle Park, NC 27709; e-mail: chris.stenland.b@bayer.com.

Abstract

BACKGROUND: Therapeutic proteins derived from human plasma and other biologic sources have demonstrated an excellent safety record relative to the potential threat of transmissible spongiform encephalopathy (TSE) transmission. Previously, hamster-adapted scrapie was used as a model agent to assess TSE clearance in purification steps leading to the isolation of biopharmaceutical proteins. The current study investigated the validity of hamster scrapie as a model for human TSE clearance studies. The partitioning of the pathogenic forms of the prion protein associated with human variant CJD (PrPvCJD), human sporadic CJD (PrPsCJD) and Gerstmann-Sträussler-Scheinker (PrPGSS) syndrome was compared to the partitioning of hamster scrapie (PrPSc) in three plasma protein purification steps. Sheep scrapie (PrPSc) was similarly evaluated.

STUDY DESIGN AND METHODS: The starting materials for three plasma protein purification steps, cryoseparation, 3 percent PEG separation, and 11.5 percent PEG separation, were spiked with brain homogenates containing human PrPvCJD, human PrPsCJD, human PrPGSS, sheep PrPSc, and hamster 263K PrPSc. The partitioning of the pathogenic form of the PrP was analyzed.

RESULTS: Clearance of the pathogenic form of the PrP was measured relative to the effluent fraction. Regardless of the source of the pathogenic prion, clearance was similar to hamster PrPSc. A nominal amount of clearance (approx., 1 log), an intermediate amount of clearance (approx., 2 log), and a substantial amount of clearance (≥3 log) were observed for the cryoseparation, 3 percent PEG separation, and 11.5 percent PEG separation steps, respectively. In the latter step, no PrP was detected in the effluents.

CONCLUSIONS: These data demonstrate that human prions, including vCJD prions, can be removed during the purification of human therapeutic proteins and indicate that partitioning of human prions is similar to that observed in the hamster scrapie model.

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