Development of analytical methods for quality control of therapeutic proteins using Quality by Design principles: the Infliximab case study

Recombinant proteins are complex molecules expressed by living organisms that present numerous possible conformations, post-translational modifications and microheterogeneity. Chemical and enzymatic modifications can readily occur during production, extraction, purification, formulation and storage. Among these modifications, some of them are particularly critical for the efficacy and safety and are known as Critical Quality Attributes (CQAs), which affect the Quality Target Product Profile (QTPP). Quality Control (QC) is a key factor in biopharmaceutical development since it allows to assess the quality of the final product and thus its safety and efficacy. It involves a series of analytical procedures designed and validated to monitor the product's CQAs. However, the complexity of recombinant proteins makes analytical method development a challenging task. Analytical Quality by Design (AQbD) could represent a structured approach to analytical method development that can help to ensure a robust procedure across its lifecycle. It consists in a systematic approach to procedure development and validation that begins with predefined objectives and emphasizes procedure performance understanding and control, based on sound science and Quality Risk Management. The QC of therapeutic monoclonal antibody Infliximab by using AQbD approach will be presented as guiding example to obtain high performance analytical procedures. Preliminary results of the set-up analytical platform development will be shown. Liquid chromatography, capillary electrophoresis and mass spectrometry were used as analytical techniques to evaluate the QTPP. The adoption of Risk Assessment facilitated the identification and management of risk factors. The application of multivariate tools, at first using a screening phase and then response surface methodology, made it possible to achieve a high degree of analytical procedure understanding.