Novel downstream processes for the purification of monoclonal antibodies based on aqueous two phase partitioning and hydrophobic interaction chromatography

Monoclonal antibodies have proven their potential as therapeutic molecules with applications as diverse as diagnosis, oncology and treatment of autoimmune diseases; the need for their supply is higher than it ever was, with the global market for monoclonal antibodies being expected to grow to nearly $58 billion in 2016. The main challenge for this growth remains the fact these products need to be manufactured cost-effectively to meet the current demand and although productive and efficient upstream processes have been developed, the downstream processing has become the new bottleneck for monoclonal antibody production. Thus, it is crucial to improve the existing purification platforms. To address this problem several alternatives were tested in, starting from the screening of new possible aqueous two phase systems (ATPS) for affinity partitioning of monoclonal antibodies with the LYTAG ligand, followed by the intensive testing of the most promising system (PEG-ammonium sulfate) with different parameters and finally a purification process combing ATPS with PEG6000-ammonium sulfate followed by hydrophobic interaction chromatography (HIC) with different commercially available resins. The introduction of an affinity ligand aimed to improve ATPS selectivity whilst taking advantage of its inherent qualities of cost effectivity, scalability and high capacity however the tested systems were fruitless in achieving a desirable purification. The HIC was introduced to increase the selectivity, still representing an accessible substitute to the traditional protein A chromatography. The combination of a 10%PEG6000-12% Ammonium Sulfate and a chromatography with HiTrap Butyl FF achieved of the best results with extraction yields up to 99% and purities as high as 96.5%.