Development, characterization and stability of therapeutic protein co-formulations

Protein therapeutics have steadily gained importance for the targeted pharmacotherapy of severe diseases in the last decades. The specific targeting of a single antigen has proven to be a successful therapeutic concept and the combination of multiple monoclonal antibodies (mAbs) in one drug product to target multiple antigens by one medication has gained increasing interest in the last years. Despite the high number of available fixed-dose co-formulations containing small molecule drugs, only a small number of fixed-dose combinations of different protein drugs has gained access to the market. This is already indicative for the clinical, regulatory, analytical and technological challenges that can arise during the development of protein co-formulations, which will be presented in Chapter I. Based on the presented challenges that can arise in drug product development of protein co-formulations, multiple clinically established protein drugs will be co-formulated to study the arising protein interactions, their impact on the individual protein stability and the effect of important formulation parameters such as pH and ionic strength on these interactions. The aim of this thesis was to identify feasible analytical approaches and formulation strategies to characterize and stabilize co-formulations of different therapeutic proteins. In Chapter II, two model protein drugs with considerably different physicochemical properties will be co-formulated to identify formulation conditions that provide sufficient stability for both proteins. In Chapter III, biophysical methods will be applied to understand how the formulation conditions affect the stability and cross-interactions between model proteins. The feasibility of these methods for the detection of protein cross-interactions in co-formulations will be investigated. In Chapter IV, a biophysical characterization of binary mixtures of mAbs that differ in net charge, charge distribution and CDR composition will be conducted to compare the colloidal and conformational stability of the proteins alone and in combination in a wide range of formulation conditions. In Chapter V two model mAbs will be selected for a more detailed analysis of the stability of the individual mAbs in co-formulations. Different ratios of both proteins will be analyzed to differentiate between self- and cross-interactions. Stability-indicating methods for co-formulations of both mAbs will be developed and forced degradation studies will be conducted to show how the stability of the mAbs is affected in the presence of different amounts of the second mAb. In Chapter VI, the in-use stabilities of dornase alfa and r-tPa will be studied after dilution in standard saline solution. The subsequent application of both enzymes is an established clinical procedure for the treatment of pleural infections. A concurrent administration of both drugs would facilitate the clinical treatment, but the compatibility and in-use stabilities of both enzymes upon concurrent administration remains unclear. Analytical techniques will be developed to investigate if the degradation of dornase alpha or r-tPa is affected in the presence of the second protein. Finally, Chapter VII provides a summary of the presented results on the development, characterization and stability of protein co-formulations in the context of the current state of the art and an outlook on the future work on protein co-formulations.