Department of Pharmaceutical and Pharmacological Sciences, Via Marzolo, 5, 35131 Padova, Italy.
Molecular and Clinical Medicine, School of Medicine, University of Dundee Nethergate, Dundee, Scotland DD1 4HN, UK.
Eur J Pharm Biopharm. 2022 Jul;176:54-74. doi: 10.1016/j.ejpb.2022.05.005. Epub 2022 May 17.
The stability of the monoclonal antibody Ipilimumab, the active ingredient of Yervoy®, used for the treatment of different types of cancer, has been investigated. Shaking/temperature, light exposure and dilution, protein drug renowned stressors, were applied on a 30-45-day series of experiments to observe the physicochemical and biological behavior of the molecule. Ipilimumab demonstrated stability under shaking and heat up to 45 days, without any unfolding during the induced combined stressors. Under artificial sunlight, the mAb showed to be sensitive even under the minimum dose tested (720 kJ/m) with formation of aggregates, particularly when diluted in glucose solution. The light-induced soluble aggregates were higher in the case of diluted samples irradiated with much higher light doses (10460 kJ/m). The aggregation of Ipilimumab took place also by irradiating the non-diluted formulation, indicating that the excipients did not protect completely the drug from photodegradation. Amino acid oxidation and deamidation were found. Anyway, after irradiation with both light doses, soluble Ipilimumab maintained its typical β-sheets structure, and the tertiary structure was nearly maintained compared to the dark. As an additional stressor test, the effect of dilution on the formulation was monitored by using a saline solution (1 mg/mL Ipilimumab) applied during hospital infusion. After two days from dilution, the protein exhibited aggregation and chemical modifications including oxidation and deamidation. When stability conditions were compromised, the viability of human cell lines treated with the stressed formulation slight decreased suggesting low potential biological toxicity of the modified mAb. As this study has demonstrated the susceptibility of Ipilimumab to light, specific solutions, and excipients as well as the use of safe light in manufacturing, handling, and storage of this drug should be promoted. Moreover, the use of proper primary and secondary packaging should be indicated to avoid the detrimental effect of light on the mAb structure and efficacy. A detailed understanding of Ipilimumab physicochemical properties, integrity, and stability could assure the best storage and manipulation conditions for its safe and successful application in cancer therapy.
本研究考察了伊匹单抗（Yervoy®的有效成分）的稳定性，伊匹单抗是一种用于治疗不同类型癌症的单克隆抗体。摇晃/温度、光照和稀释，这些都是蛋白类药物的著名应激源，在为期 30-45 天的一系列实验中，我们观察了这些分子的物理化学和生物学行为。伊匹单抗在摇晃和高达 45 天的温度下表现出稳定性，在诱导的联合应激源下没有发生任何变性。在人工阳光下，即使在测试的最低剂量（720 kJ/m）下，单抗也表现出敏感性，特别是在葡萄糖溶液中稀释时，容易形成聚集体。在稀释样品受到更高的光剂量（10460 kJ/m）照射的情况下，光诱导的可溶性聚集体更高。非稀释制剂的照射也导致了伊匹单抗的聚集，这表明赋形剂不能完全保护药物免受光降解。发现了氨基酸氧化和脱酰胺作用。无论如何，在两种光剂量照射后，可溶性伊匹单抗保持其典型的β-折叠结构，与黑暗相比，三级结构几乎保持不变。作为额外的应激源测试，通过使用生理盐水溶液（1mg/mL 伊匹单抗）在医院输注期间监测稀释对制剂的影响。稀释两天后，蛋白质表现出聚集和化学修饰，包括氧化和脱酰胺。当稳定性条件受到损害时，用应激制剂处理的人细胞系的活力略有下降，这表明修饰后的 mAb 的潜在生物学毒性较低。由于本研究表明伊匹单抗对光、特定溶液和赋形剂敏感，以及在制造、处理和储存该药物时应推广使用安全的光，因此应推广使用安全的光。此外，应指出适当的初级和二级包装，以避免光对 mAb 结构和功效的不利影响。详细了解伊匹单抗的物理化学性质、完整性和稳定性，可以确保其在癌症治疗中的安全和成功应用的最佳储存和操作条件。
