이서현 (성신여자대학교)

고병준 (성신여자대학교)

이서현 (성신여자대학교)

한승아 (성신여자대학교)

박현진 (성신여자대학교)

고병준 (성신여자대학교)

Monoclonal antibody (mAb) therapeutics are exposed to environmental stresses such as heat during production and storage, which can compromise their structural stability. Heat stress induces protein aggregation, leading to decreased therapeutic efficacy and increased immunogenicity. Recent studies suggest that such aggregation is closely associated with specific post-translational modifications (PTMs) triggered by thermal stress.

This study investigated the effects of heat stress at 37°C, representing physiological and long-term storage conditions on both aggregation behavior and PTM alterations in the monoclonal antibody Avastin. Aggregation was quantified over time points (18 h, 24 h, 48 h, 72 h, and 96 h) using HPLC-size exclusion chromatography (SEC), and PTM profiles were analyzed by peptide mapping with LC-MS/MS.

A time-dependent increase in aggregation was evident with prolonged heat exposure. Notably, aggregated fractions exhibited higher levels of deamidation compared to the native fraction. Sequence coverage differences further indicated that heat-induced PTMs may contribute to aggregation propensity.

These findings provide insight into the mechanism of antibody aggregation under thermal stress and highlight PTM signatures as potential markers for quality assessment. This study underscores the importance of controlling PTMs during antibody production, with implications for designing more stable therapeutic antibodies and improving biopharmaceutical quality assurance.