Unchained Labs eBook - 20

Unleash your biologic * See stability in hi-def with Hunky

The aggregation pathways for Adalimumab in 10 mM acetate pH 4.0,
10 mM acetate pH 5.0 and citrate-phosphate pH 5.2 were determined
by ΔGtrend (Figure 6). ΔG increases with protein concentration for all 3
formulations. The increase indicates native state aggregation needs
to be addressed with a formulation tweak. Adalimumab in citratephosphate pH 5.2 is identified as the formulation on the right track
with the highest stability (ΔG > 7.0 kcal/mol) and the smallest change
in ΔG with concentration. The aggregation pathway information
provided by ΔGtrend lets researchers know if their sample is prone
to native state aggregation, which may be addressed through
formulation tweaks, or denatured state aggregation that deserves
closer examination.

Conclusion
Many methods for studying protein stability and aggregation are
qualitative, which can cause uncertainty. In contrast, Hunky gives you
the ΔG of a protein which is a powerful and quantitative indicator of
the amount of unfolded protein present in a sample at ambient
temperatures. Inherent in the ΔG values is information about the
likelihood and pathway of aggregation of the protein. Andrew Hagarman,
PhD from KBI Biopharma finds, "With ΔG and ΔGtrend our clients are
provided with additional confirmation of a product formulation with
long term stability." Hunky fully automates chemical denaturation
experiments from start to finish allowing researchers to make biologic
stability and aggregation decisions quantitatively with ΔG.
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Figure 6. The magnitude of change between low and high concentration
ΔG(1) measurements identified 10 mM citrate, 100 mM phosphate pH 5.2
as the most stable and least aggregation prone formulation.

References
1. Freire E, Schön A, Hutchins BM, Brown RK. 2013. Chemical denaturation as a tool in
the formulation optimization of biologics. Drug Discov Today 18(19-20): 1007-1013.
2. Schön A, Clarkson BR, Siles R, Ross P, Brown RK, Freire E. 2015. Denatured state
aggregation parameters derived from concentration dependence of protein stability.
Analytical Biochemistry 488: 45-50.

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