Process and Analytical Sciences, Biopharmaceuticals R&D, AstraZeneca, Gaithersburg, MD, 20787, USA.
Dosage Form Design & Development, Biopharmaceuticals R&D, AstraZeneca, Gaithersburg, MD, USA.
Pharm Res. 2023 Oct;40(10):2355-2370. doi: 10.1007/s11095-023-03524-x. Epub 2023 May 2.
Spray-drying is considered a promising alternative drying method to lyophilization (freeze-drying) for therapeutic proteins. Particle counts in reconstituted solutions of dried solid dosage forms of biologic drug products are closely monitored to ensure product quality. We found that high levels of particles formed after reconstitution of protein powders that had been spray-dried under suboptimal conditions.
Visible and subvisible particles were evaluated. Soluble proteins in solution before spray-drying and in the reconstituted solution of spray-dried powder were analyzed for their monomer content levels and melting temperatures. Insoluble particles were collected and analyzed by Fourier transform infrared microscopy (FTIR), and further analyzed with hydrogen-deuterium exchange (HDX).
Particles observed after reconstitution were shown not to be undissolved excipients. FTIR confirmed their identity as proteinaceous in nature. These particles were therefore considered to be insoluble protein aggregates, and HDX was applied to investigate the mechanism underlying aggregate formation. Heavy-chain complementarity-determining region 1 (CDR-1) in the aggregates showed significant protection by HDX, suggesting CDR-1 was critical for aggregate formation. In contrast, various regions became more conformationally dynamic globally, suggesting the aggregates have lost protein structural integrity and partially unfolded after spray-drying.
The spray-drying process could have disrupted the higher-order structure of proteins and exposed the hydrophobic residues in CDR-1 of the heavy chain, contributing to the formation of aggregate through hydrophobic interactions upon reconstitution of spray-dried powder. These results can contribute to efforts to design spray-dry resilient protein constructs and improve the robustness of the spray-drying process.
喷雾干燥被认为是一种有前途的替代冻干（冷冻干燥）方法，可用于治疗性蛋白质。密切监测生物药物产品干燥固体剂型复溶溶液中的颗粒计数，以确保产品质量。我们发现，在喷雾干燥条件不佳的情况下，蛋白质粉末复溶后会形成高水平的颗粒。
评估可见和亚可见颗粒。在喷雾干燥之前和喷雾干燥粉末的复溶溶液中分析可溶性蛋白质的单体含量水平和熔点。收集不溶性颗粒并用傅里叶变换红外显微镜（FTIR）进行分析，并进一步用氘氢交换（HDX）进行分析。
复溶后观察到的颗粒不是未溶解的赋形剂。FTIR 证实其本质上是蛋白质。因此，这些颗粒被认为是不溶性蛋白质聚集体，并且应用 HDX 来研究聚集体形成的机制。聚集体中的重链互补决定区 1（CDR-1）在 HDX 下显示出显著的保护，表明 CDR-1 对于聚集体形成至关重要。相比之下，各种区域在全局上变得更加构象动态，表明聚集体在喷雾干燥后失去了蛋白质结构完整性并部分展开。
喷雾干燥过程可能破坏了蛋白质的高级结构，并暴露出重链 CDR-1 中的疏水性残基，在喷雾干燥粉末复溶时通过疏水相互作用导致聚集体形成。这些结果有助于设计喷雾干燥耐受的蛋白质结构并提高喷雾干燥过程的稳健性。
