290907 Role of Intermolecular Interactions On Ultrafiltration of Pegylated Proteins

Tuesday, October 30, 2012
Hall B (Convention Center )
Krisada Ruanjaikaen, Chemical Engineering, Pennsylvania State University, University Park, PA and Andrew L. Zydney, Chemical Engineering, The Pennsylvania State University, University Park, PA

PEGylated proteins are attractive as biotherapeutics due to the increased in vivo efficacy associated with the covalent attachment of polyethylene glycol (PEG) to the base protein. Previous studies have demonstrated the feasibility of using ultrafiltration for removal of residual unreacted proteins and PEG (Molek and Zydney, 2006) as well as more highly PEGylated species (Ruanjaikaen and Zydney, 2011) from the desired singly PEGylated product. In addition, ultrafiltration/ diafiltration (UF/DF) can be used for the final formulation of PEGylated proteins to achieve the desired final product concentration and buffer conditions. The objective of this study was to evaluate the effects of intermolecular interactions, mediated primarily by the PEG, on the performance of these ultrafiltration systems.

Experimental studies were performed with α-lactalbumin PEGylated with a 20 kDa activated PEG. Ultrafiltration data were obtained over a range of PEG concentration, solution conditions, and filtrate flux using Ultracel composite regenerated cellulose membranes (both as is and chemically-modified to provide a negatively-charged membrane). Data were analyzed using a recently developed theoretical model that accounts for the increase in effective size associated with the attached PEG along with ion exclusion from the PEG layer surrounding the charged protein (Molek et al., 2010).

The transmission of the PEGylated protein was significantly greater in the presence of high PEG concentrations due to the unfavorable intermolecular interactions between the free PEG and the PEGylated protein. The magnitude of this effect was consistent with previous thermodynamic models of PEG-PEG interactions. These intermolecular interactions also reduced the selectivity in ultrafiltration processes used for purification of the singly-PEGylated product. These results provide new insights into the effects of intermolecular interactions on the design and performance of ultrafiltration processes for the concentration and purification of PEGylated proteins.

References

Molek JR, Ruanjaikaen K, Zydney AL. 2010. Effect of electrostatic interactions on transmission of PEGylated proteins through charged ultrafiltration membranes. J Membr Sci 353:60–69.

Molek JR, Zydney AL. 2007. Separation of PEGylated α-lactalbumin from unreacted precursors and byproducts using ultrafiltration. Biotechnol Prog 23:1417–1424.

Ruanjaikaen K, Zydney AL. 2011. Purification of singly PEGylated α-lactalbumin using charged ultrafiltration membranes. Biotechnol Bioeng 108:822–829.


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