Formulation of stable protein powders by supercritical fluid drying

Abstract

Protein pharmaceuticals are potent drugs for the treatment of several chronic and life-threatening diseases. However, the complex and sensitive nature of protein molecules requires special attention in the development of stable dosage forms. Developing stable aqueous protein formulations is often a problem. Dried formulations have been successfully used to overcome instability of the protein drug during shipping and long-term storage. Among the drying processes available, lyophilization, or freeze-drying, is the most commonly used technique for protein pharmaceuticals. However, lyophilization is an expensive, time- and energy-consuming process. Furthermore, some proteins are susceptible to damage during the freezing step and freeze-drying is not a method of choice if microparticulate powders with well-defined particle characteristics are needed, e.g., for pulmonary delivery. Supercritical fluid (SCF) drying has been proposed as an alternative to lyophilization. SCF drying is a fast process capable of producing microparticulate powders. In SCF drying the freezing step is bypassed, but factors like mixing, mass transport rates, droplet formation and composition of the SCF fluid can play a crucial role in drying kinetics and properties of dried products, as well as in the choice of the excipients. The aim of this thesis was to investigate the capability of SCF drying for protein stabilization. The primary goal was to gain new fundamental insights into the effect of processing and formulation parameters on the stability of proteins during and after drying from aqueous solution, as well as after storage and reconstitution. In particular, relationships between processing and formulation parameters, characteristics of the dried product and protein stability were studied to identify the critical parameters for successful protein stabilization by SCF drying. To this end, we selected readily available model proteins (lysozyme, myoglobin, and IgG) with various physicochemical characteristics. The results presented in this thesis show that SCF drying is capable of producing stable proteins embedded in an amorphous carbohydrate matrix. Some general recommendations established for protein stabilization in freeze-drying were shown to be valid for SCF drying as well. Firstly, the addition of a sugar (e.g., sucrose, trehalose) to the protein formulation prevents protein aggregation in the solid state as well as after reconstitution. Secondly, an amorphous matrix with a high glass transition temperature is preferred to avoid crystallization, which is detrimental for most proteins. Stabilizers commonly used in lyophilization yielded amorphous matrices. However, in SCF drying sucrose was more difficult to process as amorphous powder. Designing a SCF drying process to obtain a stable protein formulation turned out to be a delicate interplay between excipient selection and process optimization. SCF composition was important for successful drying. Although reported to be necessary for drying of aqueous solutions, the use of ethanol in SCF drying turned out to be detrimental for formulation stability. Unexpectedly high levels of residual ethanol had a plasticizing effect, which resulted in sugar crystallization during storage. We showed that it was possible to dry without ethanol or any other co-solvent with selected formulation and process parameters and to obtain stable dry protein formulations.