Investigating the Interaction between Excipients and Monoclonal Antibodies PGT121 and N49P9.6-FR-LS: A Comprehensive Analysis

N49P9.6-FR-LS and PGT121 are promising antibodies with
significant
therapeutic potential against HIV infection, but they are prone to
precipitation at concentrations greater than 12 to 13 mg/mL. This
study evaluates the influence of six excipientsarginine, alanine,
sucrose, trehalose, methionine, and glutamateon the biophysical
stability of antibodies. We employed a comprehensive approach, combining
computational mAb-excipient interaction analysis via the site-identification
by ligand competitive saturation (SILCS) method with extensive experimental
characterization. Our experimental matrix included viscosity measurements
across temperature gradients, particle size distribution, zeta potential,
pH value, and solution appearance, alongside a short-term stability
product study at 30 °C and 65% relative humidity, with assessments
at t0 (initial), t1 (14 days), and t2 (28 days). Results indicated that sucrose, arginine, alanine, and
trehalose provided varying degrees of stabilization for both antibodies.
Conversely, glutamate destabilized PGT121 but stabilized N49P9.6-FR-LS,
while methionine had a negative effect on N49P9.6-FR-LS but a positive
one on PGT121. SILCS-Biologics analysis suggested that stabilization
by these excipients is linked to their ability to occupy regions involved
in self-protein interactions. Debye–Hückel–Henry
charge calculations further indicated that neutral excipients like
sucrose and trehalose could alter mAb charges by affecting buffer
binding, influencing aggregation propensity. These findings offer
valuable insights for optimizing antibody formulations, ensuring enhanced
product stability and therapeutic efficacy for HIV treatment.

N49P9.6-FR-LS与PGT121是极具前景的抗HIV感染治疗性抗体，但其浓度高于12~13 mg/mL时易发生沉淀。本研究评估了6种辅料——精氨酸、丙氨酸、蔗糖、海藻糖、甲硫氨酸与谷氨酸——对抗体生物物理稳定性的影响。本研究采用综合研究策略，将基于配体竞争性饱和位点识别（site-identification by ligand competitive saturation, SILCS）方法的单克隆抗体-辅料相互作用计算分析，与大规模实验表征相结合。实验矩阵涵盖了温度梯度下的黏度测定、粒径分布、Zeta电位、pH值与溶液外观表征，同时在30℃、相对湿度65%的条件下开展短期稳定性产品考察，分别在t0（初始时刻）、t1（第14天）与t2（第28天）进行检测评估。结果表明，蔗糖、精氨酸、丙氨酸与海藻糖可对两种抗体提供不同程度的稳定效果。与之相反，谷氨酸会降低PGT121的稳定性，却能稳定N49P9.6-FR-LS；而甲硫氨酸对N49P9.6-FR-LS存在不利影响，却可对PGT121起到正向稳定作用。SILCS-Biologics分析显示，这些辅料的稳定作用与其能够占据蛋白质自身相互作用区域的能力密切相关。德拜-休克尔-亨利（Debye–Hückel–Henry）电荷计算进一步证实，蔗糖与海藻糖等中性辅料可通过影响缓冲液结合改变单克隆抗体（monoclonal antibody, mAb）的电荷，进而影响其聚集倾向。本研究结果可为优化抗体制剂配方、提升HIV治疗用抗体产品的稳定性与治疗效能提供极具价值的理论指导。