Application of Instrumental Analysis Methods to Characterisation of Monoclonal Antibody-Based Formulations

Abstract

Monoclonal antibodies (mAbs) are a dominant class of protein-based drugs used in multiple therapeutic contexts. These drugs have revolutionised the management of a number conditions and are positioned as an important strategy in tailored pharmaceutical approaches (targeted therapy) for immunomodulation and oncological management. The in vivo formulation of proteins, including antibodies, is a complex process tightly regulated through environmental factors and homeostatic mechanisms. Reproduction of these conditions is essential during ex vivo manufacture of mAbs to ensure the stability of proteins and to prevent aggregation, which may impact on the efficacy of the agent in biological tissue. Among these environmental factors, temperature pH, buffer characteristics, and the use of excipients, including salts, sugars, surfactants and amino acids are of paramount importance. The focus of the present paper was to provide an overview of contemporary literature, with a focus on key studies performed by the author and colleagues, focusing on how excipient use and instrumental monitoring of stability and aggregation in mAbs could have the potential to influence future drug design and protein formulation practice. Analysis of the data set suggested that the regulation of these features during the formulation of mAbs is a complex process and there is a need to ensure that all processes within the manufacture of mAbs are controlled according to the relevant environmental factors influencing protein stability and aggregation. This paper highlights the use of adherent and non-adherent cell lines to investigate the ability of excipients to maintain mAb stability. The specific excipients analysed including amino acids (arginine and glutamate), and combinations/comparison of ionic excipients in solution, pH and dynamics of solutions, including protein-protein interactions, concentrations and agglomeration. It was demonstrated in these studies that excipients have specific and contrasting effects on mAb stability and aggregation potential. Furthermore, characterisation of the effects of excipients and formulation techniques provided a key insight into the role of these factors in maintaining mAb stability, while instrumental techniques were evaluated during the characterisation of stability and agglomeration in a variety of contexts. The majority of this paper reflects the culmination of six published papers, in which the author has actively participated, focusing on the application of instrumental analysis methods to the characterisation of mAb formulations. These studies highlight a broad range of techniques used to characterise protein formulations and which may be applied to mAbs, including nuclear magnetic resonance (NMR) spectroscopy and orthogonal techniques. Characterisation of the size of protein/mAb aggregates has been demonstrated, with pharmaceutical relevance, including in the design of pro-drugs. Furthermore, key characteristics of mAb formulation, stability and aggregation have been characterised using multiple models, providing a comprehensive insight into this topic. The included studies indicate that additives that disrupt protein-protein interactions and prevent aggregation can influence mAb stability and that the stabilisation characteristics of these proteins can be measured using instrumental techniques. These studies have clear implications for the use of instrumental analysis in the development of therapeutic mAbs, highlighting the need for further research in this field, including further investigation of the value of NMR spectroscopy and orthogonal techniques