Excipient combinations to manage protein viscosity for highly concentrated formulations

Purpose: When delivered via the subcutaneous route of application the volume of a drug is restricted to approx. 2 mL. For therapeutic proteins, such as mAbs or plasma proteins, this restriction requires the use of high protein concentrations. As can be seen by Figure 1 some proteins begin to become highly viscous at concentrations as low as 100 mg/mL. At such concentrations the formulation could potentially not even be forced through a syringe any longer. We investigate the effect of excipient combinations to manage protein viscosity.

Objectives: To provide best in class excipients to reduce viscosity and enable patient friendly administration.

Methods: Monoclonal antibodies were obtained in their respective FDA or EMA registered formulation.

For excipient studies, a chimeric monoclonal anti-TNF-α antibody at pH 7.2 was selected (mAbC). All excipients & buffer components were purchased from MilliporeSigma.

For buffer exchange and achieving high protein concentrations we used Amicon® Ultra-4 Ultracell-30k centrifugal filter units. For excipient testing five diavolumes were exchanged via centrifugation at 2,000 xg with these spin columns. Volume was also reduced by centrifugation at 2,000 xg. Protein concentration was determined according to Lambert-Beer’s law and extinction measurement at 280 nm in a BioSpectrometer® Kinetic (Eppendorf, Hamburg, Germany). Dilutions were prepared with the respective buffer and concentration verified again by the same method.

Viscosity was measured after equilibrating samples to 20 °C using a m-VROC™ viscometer at a shear rate of 1,000–3,000 /s. 200 μL of sample were loaded into a 500 μL gastight syringe (Hamilton, Reno, USA) and measured in triplicates after a priming step.

Particle diffusion Dt was measured on a Dynapro PRIII (Wyatt Technology, Santa Barbara, USA) using Dynamic Light Scattering (DLS) with 10 acquisitions of 5 seconds each at 25 °C.

The diffusion at infinite dilution D0 was derived from the equation Dt = D0 ( 1+ kD*c) by fitting the diffusion linearly over a range of 3 to 14 mg/mL mAbC. To determine the diffusion interaction parameter kD a normalized form given by Dt/D0 was plotted.

The syringe glide force is calculated according to the following equation:

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