Robustness of nanofiltration for increasing the viral safety margin of biological products.

In this study, the virus-removal capacity of nanofiltration was assessed using validated laboratory scale models on a wide range of viruses (pseudorabies virus; human immunodeficiency virus; bovine viral diarrhea virus; West Nile virus; hepatitis A virus; murine encephalomyocarditis virus; and porcine parvovirus) with sizes from 18 nm to 200 nm and applying the different process conditions existing in a number of Grifols' plasma-derived manufacturing processes (thrombin, α1-proteinase inhibitor, Factor IX, antithrombin, plasmin, intravenous immunoglobulin, and fibrinogen). Spiking experiments (n = 133) were performed in process intermediate products, and removal was subsequently determined by infectivity titration. Reduction Factor (RF) was calculated by comparing the virus load before and after nanofiltration under each product purification condition. In all experiments, the RFs were close to or greater than 4 log10 (>99.99% of virus elimination). RF values were not significantly affected by the process conditions within the limits assayed (pH, ionic strength, temperature, filtration ratio, and protein concentration). The virus-removal capacity of nanofiltration correlated only with the size of the removed agent. In conclusion, nanofiltration, as used in the manufacturing of several Grifols' products, is consistent, robust, and not significantly affected by process conditions.

Capacity of the manufacturing process of Flebogamma(®) DIF, a new human high purity intravenous immunoglobulin, to remove a TSE model-agent.

The variant Creutfeldt-Jakob disease (vCJD) is a transmissible spongiform encephalopathy (TSE) associated with the ingestion of cattle derived products affected with bovine spongiform encephalopathy. vCJD emerged in the UK, where most of the cases occurred (170 of 217 cases worldwide). Manufacturers of biological products must investigate the ability of their production processes to remove TSE agents. Two manufacturing steps (polyethylene glycol-PEG precipitation and nanofiltration down to 20 nm) of Flebogamma(®) DIF, were evaluated by western blot and bioassay to measure the prion protein (PrP(Sc)) and infectivity clearance capacity, respectively. A laboratory scale model representative of the industrial process and a (experimentally) spiked TSE model-agent (hamster scrapie strain 263 K) were employed. Both steps showed a significant capacity to clear the TSE model-agent used since no PrP(Sc) signal or infectivity was detected in the resulting product of each step. PEG precipitation and nanofiltration provided reduction factors of ≥6.19 log(10)ID(50) and ≥5.45 log(10)ID(50) respectively. Both steps showed consistency between western blot and bioassay results. These results demonstrate the ability of the Flebogamma(®) DIF manufacturing process to clear TSE agents beyond the limit of detection of the assays, by several orders of magnitude.

A prospective study comparing human metapneumovirus with other respiratory viruses in adults with hematologic malignancies and respiratory tract infections.

Human metapneumovirus (hMPV) is a recently described paramyxovirus associated with upper and lower respiratory-tract infection (URI and LRI, respectively). We conducted a prospective study of URI and LRI in adults with hematologic malignancies during a 4-year period. We retrospectively tested samples by reverse-transcription polymerase chain reaction for hMPV and analyzed clinical data. Twenty-two (9%) of 251 episodes of respiratory infection tested positive for hMPV. Sixteen (73%) of the illnesses occurred in hematopoietic stem-cell transplant recipients. Nine patients with hMPV developed LRI; 3 of these patients died. hMPV is a common cause of respiratory infections in adults with hematologic malignancies, with associated morbidity and mortality.