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Pharmaceutical Research

, Volume 30, Issue 4, pp 968–984 | Cite as

Effect of pH and Excipients on Structure, Dynamics, and Long-Term Stability of a Model IgG1 Monoclonal Antibody upon Freeze-Drying

  • Jihea Park
  • Karthik Nagapudi
  • Camille Vergara
  • Ranjini Ramachander
  • Jennifer S. Laurence
  • Sampathkumar KrishnanEmail author
Research Paper

Abstract

Purpose

To investigate the mechanism of IgG1 mAb stabilization after freeze-drying and the interdependence of protein structural preservation in the solid state, glassy state dynamics and long-term storage stability under different formulation conditions.

Methods

IgG1 mAb was formulated with mannitol at pH 3.0, 5.0, and 7.0 in the presence and absence of sucrose and stability was monitored over 1 year at different temperatures. Physical and covalent degradation of lyophilized formulation was monitored using SEC, CEX, and light obscuration technique. Secondary and tertiary structure of the protein in the solid state was characterized using FTIR and fluorescence spectroscopy respectively. Raman spectroscopy was also used to monitor changes in secondary and tertiary structure, while SS-NMR 1H relaxation was used to monitor glassy state dynamics.

Results

IgG1 mAb underwent significant secondary structural perturbations at pH 3.0 and conditions without sucrose, while pH 5.0 condition with sucrose showed the least structural change over time. The structural changes correlated with long-term stability with respect to protein aggregate formation and SbVP counts. SS-NMR data showed reduced relaxation time at conditions that were more stable.

Conclusions

Native state protein structural preservation and optimal solid-state dynamics correlate with improved long-term stability of the mAb in the different lyophilized formulations.

KEY WORDS

IgG1 antibody lyophilization protein formulation protein structure solid state NMR solid state stability 

Abbreviations

CEX

cation-exchange chromatography

FTIR

fourier transform infrared spectroscopy

HMWS

high molecular weight species

HPLC

high performance liquid chromatography

LMWS

low molecular weight species

mAb

monoclonal antibody

RWC

residual water content

SbVP

sub-visible particle

SEC

size-exclusion chromatography

SS-NMR

solid state nuclear magnetic resonance.

Tg

glass transition temperature

XRD

X-ray diffraction

Notes

Acknowledgments and Disclosures

Thanks to Twinkle Christian and Chris Woods for helping with FTIR analysis. Special thanks to Dr. Margaret Ricci, Dr. Ping Yeh, Prof. David Volkin and Dr. David Brems for their technical input and support.

Supplementary material

11095_2012_933_MOESM1_ESM.docx (152 kb)
ESM 1 (DOCX 151 kb)

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Copyright information

© Springer Science+Business Media New York 2012

Authors and Affiliations

  • Jihea Park
    • 1
  • Karthik Nagapudi
    • 2
  • Camille Vergara
    • 1
  • Ranjini Ramachander
    • 1
  • Jennifer S. Laurence
    • 3
  • Sampathkumar Krishnan
    • 1
    Email author
  1. 1.Amgen Inc.Process and Product DevelopmentThousand OaksUSA
  2. 2.Amgen Inc.Small Molecule Process and Product DevelopmentThousand OaksUSA
  3. 3.Department of Pharmaceutical ChemistryUniversity of KansasLawrenceUSA

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