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FTIR Spectroscopy Detects Intermolecular β-Sheet Formation Above the High Temperature Tm for Two Monoclonal
Garrett Baird1, Chris Farrell2, Jason Cheung3
1Merck & Co., Inc., MMD, West Point, PA, USA.
The Protein Journal
|July 14, 2020
Summary
Fourier Transform Infrared (FTIR) spectroscopy revealed that monoclonal antibodies undergo structural changes from intramolecular to intermolecular beta sheets at higher temperatures, leading to aggregation. Sucrose concentration impacts this antibody stability.
Area of Science:
- Biochemistry
- Spectroscopy
- Protein Chemistry
Background:
- Monoclonal antibodies (mAbs) are crucial therapeutics, but their stability during formulation is critical.
- Understanding antibody conformational changes and aggregation is essential for developing effective drug products.
Purpose of the Study:
- To investigate the temperature-dependent secondary structure changes of two IgG monoclonal antibodies (anti-IGF1R and anti-TSLP).
- To elucidate the impact of formulation conditions, specifically sucrose concentration, on antibody stability and aggregation.
Main Methods:
- Transmission mode Fourier Transform Infrared (FTIR) spectroscopy, including second derivative analysis.
- Differential Scanning Calorimetry (DSC) to determine thermal transition temperatures (Tm1 and Tm2).
- Thermal Scanning Dynamic Light Scattering (TS-DLS) to assess particle size changes and aggregation.
Main Results:
- Both antibodies exhibited two major thermal transitions (Tm1 near 70°C, Tm2 near 85°C) via DSC.
- TS-DLS indicated significant particle size increase and aggregation near Tm2.
- FTIR spectroscopy clearly identified a transition from intramolecular to intermolecular beta-sheet structures in both antibodies with increasing temperature.
- FTIR also demonstrated the influence of sucrose concentration on antibody secondary structure and stability.
Conclusions:
- FTIR spectroscopy is a powerful tool for studying temperature-induced secondary structure changes and aggregation in monoclonal antibodies.
- The study identified specific structural transitions associated with antibody aggregation, providing insights for formulation development.
- Excipients like sucrose play a significant role in maintaining antibody stability, as evidenced by FTIR analysis.
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