CJD PrP sc removal by nanofiltration process: Application to a therapeutic immunoglobulin solution (Lymphoglobuline ®)

Abstract

The characteristic of transmissible spongiform encephalopathies (TSE) is an accumulation of partially protease resistant (PrP res) abnormal prion protein (PrP sc). This pathological prion protein is very resistant to conventional inactivation methods. The risk of transmission of TSE, such as Creutzfeldt–Jakob disease (CJD), by biopharmaceutical products prepared from human cells must be taken into account. The nanofiltration process has been proved to be effective in removing viruses and scrapie agent. The major advantages of this technique are flexibility and efficacy in removing infectious particles without altering biopharmaceutical characteristics and properties. This study focused on the removal of human PrP sc by means of a nanofiltration method after spiking a Lymphoglobuline ® solution with a CJD brain homogenate. Lymphoglobuline ® equine anti-human thymocyte immunoglobulin is a selective immunosuppressive agent acting mainly on human T lymphocytes. The therapeutic indications are: • immunosuppression for transplantation: prevention and treatment of graft rejection; • treatment of aplastic anemia. In our study, CJD homogenate was spiked at three different dilutions (low, moderate and high) in the Lymphoglobuline ® product. The nanofiltration process was performed on each sample. Using the western blot technique, the PrP res signal detected in nanofiltrates was compared to that obtained with a reference scale (dilution series of CJD brain homogenate in Lymphoglobuline ® detected by western blot and elaborated on 3.3 log). After nanofiltration, the PrP res western blot signal was detected with a significant reduction in the less dilute sample, whereas the signal was undetectable in the two other samples. These are the first data in CJD demonstrating a clearance between 1.6 and 3.3 log with a Lymphoglobuline ® recovery of over 93%. The nanofiltration process confirms its relative efficacy in removing human CJD PrP sc.