Department of Chemical and Biomolecular Engineering, and Center for Molecular Thermodynamics, University of Delaware, Newark, Delaware 19716, USA.
J Pharm Sci. 2012 Oct;101(10):3651-60. doi: 10.1002/jps.23264. Epub 2012 Jul 17.
Visible and subvisible particle formation during the storage of protein solutions is of increasing concern for pharmaceutical products. Previous work (Li Y, Ogunnaike BA, Roberts CJ. 2010. J Pharm Sci 99:645-662) showed that the model protein, alpha-chymotrypsinogen A (aCgn), forms non-native aggregates under accelerated (heated) conditions, but the size and morphology of the resulting aggregates depended sensitively on pH and NaCl. Here, it is shown that aggregates created as high-molecular-weight soluble aggregates undergo a pH- and salt-dependent reversible phase transition to a condensed or insoluble phase of suspended microparticles, whereas monomers remain completely soluble in the same regime. The location of the phase boundary is quantitatively consistent with the different regimes of kinetic behavior observed previously for aCgn. This suggests that the while kinetics is important for controlling the rates of monomer loss during non-native aggregation, it may be possible to tune solution thermodynamics and phase behavior to suppress otherwise soluble aggregates from propagating to form visible or large subvisible particles. Interestingly, the aggregate phase boundary is sensitive to the identity of salt anions in solution, highlighting the importance of electrostatics and preferential salt interactions in mediating aggregate condensation and particle formation.
在蛋白质溶液储存过程中可见和亚可见颗粒的形成引起了人们对药物产品越来越多的关注。先前的工作（Li Y, Ogunnaike BA, Roberts CJ. 2010. J Pharm Sci 99:645-662）表明，模型蛋白α-糜蛋白酶原 A（aCgn）在加速（加热）条件下形成非天然聚集物，但聚集物的大小和形态对 pH 和 NaCl 敏感依赖。在这里，研究表明形成的高分子量可溶性聚集物经历了一个 pH 和盐依赖性的可逆相转变，从可溶的凝聚或不溶的悬浮微粒子相转变，而单体在相同条件下仍完全可溶。相边界的位置与先前观察到的 aCgn 不同动力学行为的不同区域定量一致。这表明，虽然动力学对于控制非天然聚集过程中单体损失的速率很重要，但可能可以调节溶液热力学和相行为，以抑制原本可溶的聚集物进一步传播形成可见或较大的亚可见颗粒。有趣的是，聚集相边界对溶液中盐阴离子的特性敏感，突出了静电作用和优先盐相互作用在调节聚集物凝聚和颗粒形成中的重要性。
