Pharmaceutical Development, Genentech Inc. South San Francisco, South San Francisco, California, 94080, USA.
Pharm Res. 2020 Apr 26;37(5):85. doi: 10.1007/s11095-020-02798-9.
To evaluate the stability of a model Mab1 and Fab1 under in vitro vitreal conditions in the presence of various metabolites and in the presence of light at λ > 400 nM.
A full length IgG1 monoclonal antibody (Mab1) and a fab fragment (Fab1) were formulated with various metabolites typically found in the vitreous humor and subjected to visible light treatment. Both proteins were analyzed using a variety of biochemical techniques. Furthermore, Fab1 was also tested for antigen binding ability using surface plasmon resonance.
Our data shows that some vitreal metabolites such as riboflavin and ascorbic acid affect protein stability, via formation of reactive oxygen species (ROS) and advanced glycation end products (AGE) s respectively. In contrast, metabolites such as glutathione may protect these proteins from light-induced degradation to some extent.
Ascorbic acid and riboflavin were found to photosensitize therapeutic proteins especially when exposed to light. Ascorbic acid reacted with proteins even in the absence of light. Antioxidants such as glutathione helped limit photooxidation under ambient or blue light exposure. Since antioxidant capacity in the eye decreases with age we recommend understanding long term stability, including photooxidation and photosensitization, of new candidate proteins in the context of controlled or sustained release technologies for ocular diseases.
评估模型 Mab1 和 Fab1 在存在各种代谢物和 λ>400nm 光的情况下在玻璃体条件下的稳定性。
全长 IgG1 单克隆抗体（Mab1）和 Fab 片段（Fab1）与通常存在于玻璃体液中的各种代谢物一起配制，并进行可见光处理。使用各种生化技术分析两种蛋白质。此外，还使用表面等离子体共振法测试 Fab1 的抗原结合能力。
我们的数据表明，一些玻璃体液代谢物，如核黄素和抗坏血酸，分别通过形成活性氧（ROS）和晚期糖基化终产物（AGE）s 来影响蛋白质稳定性。相比之下，谷胱甘肽等代谢物可能在一定程度上保护这些蛋白质免受光诱导的降解。
抗坏血酸和核黄素被发现会使治疗蛋白敏化，尤其是在暴露于光下时。抗坏血酸即使在没有光的情况下也会与蛋白质发生反应。抗氧化剂如谷胱甘肽有助于在环境或蓝光照射下限制光氧化。由于眼睛中的抗氧化能力随着年龄的增长而下降，因此我们建议在控制或持续释放技术的背景下了解新候选蛋白的长期稳定性，包括光氧化和光敏化，以治疗眼部疾病。
