Second-generation nanofiltered plasma-derived mannan-binding lectin product: process and characteristics.

Area of Science:

• Immunology
• Biochemistry
• Protein Chemistry

Background:

• Mannan-binding lectin (MBL) is crucial for innate immunity, aiding pathogen recognition and complement activation.
• Low MBL levels increase susceptibility to severe infections, making MBL substitution therapy a promising treatment.
• A first-generation MBL product demonstrated safety and patient benefit.

Purpose of the Study:

• To develop a nanofiltered second-generation MBL product with enhanced safety and efficacy.
• To optimize MBL purification using a novel affinity matrix.
• To characterize the improved MBL product.

Main Methods:

• Screening of carbohydrate-based gels for affinity matrices.
• MBL extraction from fraction III and purification using Superdex 200 pg.
• Two virus reduction steps: Planova 20N nanofiltration and solvent/detergent treatment.
• Further purification by anion-exchange and gel-filtration chromatography.

Main Results:

• Superdex 200 pg proved superior for MBL affinity purification from fraction III, achieving a 70% yield.
• Nanofiltration through Planova 20N significantly increased viral safety with minimal MBL loss.
• The final MBL fraction (53% purity) contained high-oligomeric MBL, associated proteases (MASP), and a minor protein impurity (alpha2-macroglobulin).
• Functional activity (mannan-binding, opsonic) remained intact, but C4 activation capacity was reduced by half.

Conclusions:

• A second-generation MBL production process was successfully developed, yielding an average of 50%.
• Nanofiltration of MBL-MASP complexes through Planova 20N enhanced product safety with minimal MBL loss.