Advances in HPLC & Chromatography Techniques

Biography

Biography: Gaelle Coussot

Abstract

Monoclonal antibodies (mAb) represent the largest class of therapeutic molecules entering clinical studies. Due to their inherent structural complexity, the quality control (QC) of such type of molecules, necessary for their development and commercialization, represents a challenging analytical task. Common approaches in mAb QC are based on mAb reduction or proteolysis to produce a mixture of mAb fragments that are further analyzed by separation methods. These cleavage steps are usually performed off line (i.e. prior to the separation step in a distinct reactor). This is an important limitation in terms of time, reactant consumption and cross contamination by the possible formation of endogenous compounds that may further impair the quality assessment of the mAb drug. To overcome these limitations, we have first developed a fully integrated bio-analytical miniaturized methodology called D-PES (Diffusion–mediated Proteolysis and Electrophoretic Separation) for the QC of polymer-drug conjugates. With the D-PES methodology, both cleavages and separation steps are performed in-line, silica capillary being used both as nano-reactor and separation support. The methodology is based on transverse diffusion of laminar flow profile (TDLFP) mixing of reactant nano-volumes (proteolytic buffer (PB), substrate (S), enzyme (E) or reducing agent (R)) inside the capillary. Principles and results of these rapid and low operating costs microanalyses will be presented for mAb drugs. Separation optimization and mAb cleavage conditions (choice of background electrolyte, PB, ionic strength, pH…) will be discussed to demonstrate the robustness of the D-PES methodology.