Two-Step Size-Exclusion Nanofiltration of Prothrombin Complex Concentrate Using Nanocellulose-Based Filter Paper

• Main Authors: Levon Manukyan, Athanasios Mantas, Mikhail Razumikhin, Andrey Katalevsky, Eugen Golubev, Albert Mihranyan
• Format: Article
• Language: English
• Published: MDPI AG 2020-03-01
• Series: Biomedicines
• Subjects: mille-feuille filter; cladophora cellulose; hemophilia b; protein aggregates; virus removal filtration
• Online Access: https://www.mdpi.com/2227-9059/8/4/69

<b> </b>Coagulation Factor IX-rich protrhombin complex concentrate (FIX-PCC) is a therapeutic biologic product that consists of a mixture of several human plasma-derived proteins, useful for treating hemophilia B. Due to its complex composition, FIX-PCC is very challenging to bioprocess through virus removing nanofilters in order to ensure its biosafety. This article describes a two-step filtration process of FIX-PCC using a nanocellulose-based filter paper with tailored porosity. The filters were characterized with scanning electron microscopy (SEM), cryoporometry with differential scanning calorimetry, and nitrogen gas sorption. Furthermore, in order to probe the filter&#8217;s cut-off size rejection threshold, removal of small- and large-size model viruses, i.e., &#934;X174 (28 nm) and PR772 (70 nm), was evaluated. The feed, pre-filtrate, and permeate solutions were characterized with mass-spectrometric proteomic analysis, dynamic light scattering (DLS), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and analytical size-exclusion high-performance liquid chromatography (SEHPLC). By sequential filtration through 11 &#956;m pre-filter and 33 &#956;m virus removal filter paper, it was possible to achieve high product throughput and high virus removal capacity. The presented approach could potentially be applied for bioprocessing other protein-based drugs.