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Analytical Techniques for Evaluating Protein Instability at Interfaces

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Protein Instability at Interfaces During Drug Product Development

Part of the book series: AAPS Advances in the Pharmaceutical Sciences Series ((AAPS,volume 43))

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Abstract

Proteins at interfaces present a number of analytical challenges, particularly in the context of biotherapeutic formulations. Proteins typically adsorb to interfaces in thin films or monolayers while being present at high concentrations in the bulk liquid, making sensitivity to behavior directly at the interface an important feature. Additionally, there are multiple timescales of interest, from milliseconds to days, meaning the analytical techniques for probing interfacial behavior need to be adaptable with respect to time resolution. Also, biotherapeutic formulations are usually multicomponent systems, so the techniques need to distinguish between the behavior of the therapeutic protein and other surface-active excipients included to stabilize the protein (e.g., polysorbates and poloxamers). Finally, proteins form amorphous films at interfaces that make traditional interfacial techniques, usually applied to lipids and other well-defined surfactants, less sensitive to structural changes in protein films.

In this chapter, we discuss the strengths and weaknesses of a multitude of interfacial techniques and how these have been applied to understand protein instability at interfaces. First, we examine the multitude of techniques for measuring protein adsorption to interfaces including surface tensiometry, microscopy, and interferometry. Next, we give an overview of rheological tools for evaluating interfacial stresses in protein films. Finally, we discuss a number of spectroscopic techniques for measuring protein structure and conformation within interfacial films. In each section, we review how these various techniques have been employed to understand the dynamics of protein adsorption, unfolding, and aggregation at interfaces and how they could aid in reducing the impact of interfacial stresses during the development of biotherapeutics.

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Abbreviations

AFM:

Atomic force microscopy

ATR-FTIR:

Attenuated total reflection Fourier transform infrared spectroscopy

BAM:

Brewster angle microscopy

BLI:

Bio-layer interferometry

DPI:

Dual polarization interferometry

FRAP:

Fluorescence recovery after photobleaching

FRET:

Förster resonance energy transfer

IgG:

Immunoglobulin G

IRRAS:

Infrared reflectance absorption spectroscopy

IRSE:

Infrared spectroscopic ellipsometry

IV:

Intravenous

mAb:

Monoclonal antibody

NR:

Neutron reflection

OWLS:

Optical waveguide lightmode spectroscopy

PM:

Polarization modulated

Q:

Wave vector transfer

QCM-D:

Quartz crystal microbalance with dissipation monitoring

SFG:

Sum frequency generation

SPR:

Surface plasmon resonance

TIRF:

Total internal reflection fluorescence

XRR:

X-ray reflectivity

Δ:

Wave phase angle

Ψ:

Wave amplitude difference

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Authors and Affiliations

  1. Pharmaceutical Development, Genentech, Inc., South San Francisco, CA, USA

    Ian C. Shieh

  2. Formulation Development, Regeneron Pharmaceuticals, Inc., Tarrytown, NY, USA

    Yuan Cheng

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  1. Ian C. Shieh
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  2. Yuan Cheng
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Correspondence to Ian C. Shieh .

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Editors and Affiliations

  1. Department of Formulation Development Nevakar Inc., Bridgewater, NJ, USA

    Jinjiang Li

  2. Drug Product Development, Bristol Myers Squibb, New Brunswick, NJ, USA

    Mary E. Krause

  3. Chemical Engineering, The City College of New York - CUNY, New York, NY, USA

    Raymond Tu

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Shieh, I.C., Cheng, Y. (2021). Analytical Techniques for Evaluating Protein Instability at Interfaces. In: Li, J., Krause, M.E., Tu, R. (eds) Protein Instability at Interfaces During Drug Product Development. AAPS Advances in the Pharmaceutical Sciences Series, vol 43. Springer, Cham. https://doi.org/10.1007/978-3-030-57177-1_7

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