Mechanism of a decrease in potency for the recombinant influenza A virus hemagglutinin H3 antigen during storage.

Area of Science:

• Vaccinology
• Protein Biochemistry
• Biophysical Chemistry

Background:

• Flublok®, a recombinant hemagglutinin (rHA)-based influenza vaccine, offers an alternative to traditional egg-derived vaccines.
• This vaccine contains threefold higher hemagglutinin content compared to standard inactivated influenza vaccines.

Purpose of the Study:

• To investigate the causes and mechanisms behind the observed in vitro potency loss of rHA.
• To evaluate the biophysical and chemical stability of rHA under various formulations and storage conditions.

Main Methods:

• Characterization of rHA protein antigen.
• Assessment of higher order structure and hydrodynamic size.
• Measurement of free thiol content and disulfide-linked multimer formation.

Main Results:

• A rapid loss of in vitro potency (∼50%) was observed in rHA after 1 month of storage.
• Potency loss did not correlate with changes in higher order structure or hydrodynamic size.
• Disulfide-mediated cross-linking, forming non-native multimers, correlated with potency loss.

Conclusions:

• The primary mechanism for early potency loss in rHA is likely disulfide-mediated cross-linking.
• Loss of free thiol content, especially in C-terminal cysteine residues, is associated with decreased SRID potency.