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Related Experiment Videos

Formulation development and primary degradation pathways for recombinant human nerve growth factor

M Eng1, V Ling, J A Briggs

  • 1Department of Medicinal and Analytical Chemistry, Genentech, Inc., South San Francisco, California 94080, USA.

Analytical Chemistry
|October 24, 1997
PubMed
Summary

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Recombinant human nerve growth factor (NGF) is most stable in pH 5.5 acetate buffer at 5°C for 1.5 years. Key degradation pathways include Asp60-Pro61 cleavage, oxidation, deamidation, and Asp93 isomerization.

Area of Science:

  • Biochemistry
  • Protein Chemistry
  • Pharmaceutical Stability

Background:

  • Recombinant human nerve growth factor (NGF) is crucial for neuronal survival and function.
  • Understanding NGF stability is essential for developing effective therapeutic formulations.
  • Previous studies have highlighted the sensitivity of proteins to pH and temperature variations.

Purpose of the Study:

  • To investigate the chemical and physical stabilities of recombinant human NGF in aqueous solutions.
  • To identify degradation pathways and products under various storage conditions.
  • To determine optimal formulation conditions for long-term NGF storage.

Main Methods:

  • NGF stability was assessed at temperatures ranging from 5 to 37°C and pH levels from 4.2 to 5.8.

Related Experiment Videos

  • Degradation products were analyzed using cation-exchange chromatography, RP-HPLC, electrospray mass spectrometry, peptide mapping, and LC/MS.
  • Accelerated degradation studies were performed using base, hydrogen peroxide, and elevated temperatures.

    • Main Results:

    • NGF chemical stability decreased with decreasing pH, primarily due to Asp60-Pro61 cleavage.
    • Aggregation was a significant degradation pathway at 37°C, particularly in succinate buffer.
    • Identified degradation products included mono- and di-oxidized NGF (Met37, Met92), deamidated NGF (Asn45), and isomerized NGF (Asp93).
    • NGF in pH 5.5 acetate buffer at 5°C showed <10% degradation over 1.5 years, with Asp93 isomerization being the main pathway.

    Conclusions:

    • NGF exhibits optimal stability in pH 5.5 acetate buffer at 5°C, allowing for extended storage periods.
    • Understanding specific degradation pathways like Asp60-Pro61 cleavage and Asp93 isomerization is key to improving NGF formulations.
    • The characterized degradation products and analytical methods provide a foundation for future quality control of NGF therapeutics.