White paper: Viscosity reducing excipients for protein formulation
Protein viscosity is one of the major obstacles in preparing highly concentrated protein formulations suitable for subcutaneous (subQ) injection. Highly viscous protein solutions would require a significant force to be applied to the syringe for injection. As a result, the patient could experience a considerable amount of pain. In many cases, injectability would not be possible.1,2 When characterizing protein viscosity behavior, one can differentiate two different concentration regimes as shown in Figure 1. At very low concentrations below about 75 mg/mL, proteins are rarely viscous. When increasing the concentration to between 100 and 200 mg/mL, some proteins exhibit elevated viscosity exceeding the limit of injectability, which is typically between 20 and 25 mPa·s.
At this concentration regime, several proteins exhibit an affinity for self-interaction, i.e. forming transient clusters that give rise to elevated viscosity. At concentrations above 200 mg/mL, the nearest neighbor distance between the protein molecules shrinks so that without a specific affinity for self-interactions, said protein-protein interactions take place. While viscosity-reducing excipients can affect proteins exhibiting either of these interaction patterns, they are likely to be more efficient at protein concentration regimes below 200 mg/mL.3,4 These intermolecular interactions between proteins have the same molecular origin as the intramolecular interactions that structurally stabilize the proteins. This means viscosity-reducing excipients that affect protein-protein interactions can potentially also destabilize proteins. As such, it is essential to balance an excipient’s viscosity-reducing ability against its potential to destabilize a protein. For some excipients, a concentration-dependent effect on protein stability is well-documented.
At lower concentrations, the excipients act as stabilizers, but this behavior changes as concentration increases, often with an adverse effect on protein stability. Excipient concentration is thus a critical factor in managing protein stability. These two aspects can be better balanced by using an excipient combination of an amino acid and an anionic excipient. When used in combination, excipients are more efficient in reducing viscosity and may even do so in an over-additive manner. Consequently, lower concentrations of the individual excipients can be used, which is more favorable for protein stability.
This white paper evaluates the viscosity-reducing capacities of excipients and excipient combinations. It shows the over-additive effect of using two excipients together and addresses how excipients’ viscosityreducing ability depends on pH. The results show the effect of protein viscosity on injection force and highlight the platform’s ability to balance viscosity reduction with protein stability. The case studies presented demonstrate that using a combination of two excipients at lower concentrations instead of a single excipient at a higher concentration enables balancing protein viscosity and protein stability in a favorable way.
Table 1: Excipients and abbreviations
|Thiamine phosphoric acid ester chloride dihydrate||TMP|
Table 2: Materials used for base buffer preparation
|Acetate buffer||Acetic acid (glacial) 100% EMPROVE® EXPERT Ph Eur, BP,JP,USP|
|Sodium hydroxide solution 32% EMPROVE® EXPERT|
|Phosphate buffer||Sodium dihydrogen phosphate monohydrate EMPROVE® EXPERT BP,USP|
|di-Sodium hydrogen phosphate heptahydrate EMPROVE® EXPERT DAC,USP|
|Optional addition: Sodium chloride EMPROVE® EXPERT Ph Eur,BP,ChP,JP,USP|
Authors: Stefan Braun, Niels Banik, Jennifer J. Widera, Jan Gerit Brandenburg and Tobias Rosenkranz. Merck/Millipore Sigma.