Process and product monitoring of recombinant DNA-derived biopharmaceuticals with high-performance capillary electrophoresis.

Abstract

High-performance capillary electrophoresis (HPCE) has emerged over the past 20 years as a powerful multidimensional separation tool that is orthogonal to HPLC and comparable to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) slab gel methods. HPCE is most frequently applied in the QC release testing of recombinant DNA-derived protein and monoclonal antibody (MAb) biopharmaceuticals. HPCE is a rugged and robust separation tool that can be used like HPLC to monitor the purification process, as well as to analyze bulk drug and drug substances. Examples of the practical applications of the predominant free-solution capillary electrophoresis (FSCE) and capillary gel electrophoresis (CGE) formats of HPCE, applied for process monitoring and product monitoring of recombinant protein and MAb biotherapeutics, are presented. HPCE has been applied in FSCE mode to monitor the purification of the rDNA-derived protein, recombinant human interleukin-4 (rhIL4). FSCE is demonstrated to be a robust method that can be used to monitor multiple column chromatographic purification processes, such as immobiilized metal-ion affinity chromatography (IMAC), ion exchange chromatography (IEC), and size exclusion chromatography (SEC) columns. The FSCE data are used to pool fractions to carry forward for further purification. The FSCE method is compared to the corresponding RP-HPLC method for rhIL4. HPCE has been applied in the CGE mode to monitor the purification of an rDNA-derived IgG4 MAb. CGE is demonstrated to be a convenient and rapid method to profile the purification process, compare purification processes, and provide a fingerprint of the MAb bulk drug that is useful for determining purity and lot-to-lot consistency. The practical advantages and limitations of CGE for process monitoring and product monitoring of MAbs are presented. The CGE method is compared to the high-performance SEC separation of the MAb under nondenaturing (HP-SEC) and denaturing (HP-DSEC) conditions and to SDS-PAGE gels for the analysis and characterization of MAb bulk drugs.