High Resolution Superficially Porous Materials for Native Protein Separations

摘要 生命科学研究和其他关键的生物分析应用将大大受益于 更快、更高分辨率的高效液相色谱（HPLC）分离， 分子包括蛋白质、肽、糖肽和聚糖。蛋白质疗法很复杂 分子，易受有意或无意的组成和构象改变的影响。许多 治疗性蛋白质本质上是密切相关的变体的混合物。目前的做法是雇用 HPLC用于分离和定量蛋白质混合物等组分，以确保 蛋白质药物许多蛋白质的HPLC分离受限于蛋白质混合物的分辨率， 需要一个小时或更长时间，并且可能无法完全解决所有组件。由于复杂性 在当前的研究文献中，有许多实例将联合收割机多个HPLC 分析以获得蛋白质组成的全部细节，并测量蛋白质变体的水平， 在药物开发以及生产和质量保证/质量控制期间。 无论定量信息的预期用途如何，高效LC分离都是一种有效的方法。 分析系统的基本部分，以及实现高分辨率分离的时间 蛋白质样本是一个很大的瓶颈。该提案描述了一种改进分离的方法， 通过扩展新型表面多孔颗粒的使用提高蛋白质分析的HPLC效率 (SPP)具有优化特性的技术，提供更快、更高的分辨率 分离天然状态下的蛋白质。我们的目标是制造SPP二氧化硅颗粒， 专门设计用于保持完整天然结构的条件下，优化材料 特性，并将这些材料有效地加载到HPLC柱格式中。拟议的工作将 扩展最近在材料科学方面的突破， 材料，寻求在HPLC操作的其他原生模式中使用该技术，包括 疏水相互作用和离子交换。拟议工作的目标将产生非常高的收益， 性能色谱产品，大于目前可用的，将被广泛应用 用于蛋白质分析，特别是用于蛋白质治疗。这项技术的应用并不是 仅限于生物制药制剂，并且事实上，将有益于任何使用 用于较大生物分子分析的HPLC方法。所描述的分离技术将 直接导致有用的产品，对于这些产品存在显著的技术和市场需求。

Abstract Life sciences research and other critical bioanalytical applications would strongly benefit from faster and higher resolution high performance liquid chromatographic (HPLC) separations of larger molecules including proteins, peptides, glycopeptides, and glycans. Proteins therapeutics are complex molecules, susceptible to intended or unintended alterations of composition and conformation. Many therapeutic proteins are inherently a mixture of closely related variants. Current practice is to employ HPLC to resolve and quantify the components such protein mixtures, to assure identity and safety of the protein drug. Many HPLC separations of proteins are limited in resolution of protein mixtures, require an hour or more, and may not completely resolve all of the components. Due to the complexity of samples, there are many examples in the current research literature that combine multiple HPLC analyses to gain full details on protein composition, and to measure the levels of protein variants, both during development of the drug, as well as manufacture and quality assurance/quality control. Regardless of the intended use of the quantitative information, high efficiency LC separations are a fundamental part of the analytical systems, and the time to achieve high resolution separations of protein samples is a great bottleneck. This proposal describes an approach to improve the separation efficiency of HPLC for protein analysis by extending the use of novel superficially porous particle (SPP) technologies with optimized characteristics, providing significantly faster and higher resolution separations of proteins in the native state. Our objectives are to create SPP silica particles, specifically design for use in conditions that maintain intact native structures, optimize the materials properties, and to load these materials efficiently into HPLC column formats. The proposed work will extend recent breakthroughs in material science that yield authentic improvements in current materials, seeking to employ the technology in additional native modes of HPLC operation, including hydrophobic interaction and ion exchange. The goals of the proposed work would yield very high performance chromatographic products, greater than those currently available, to be applied broadly in analysis of proteins, particularly for protein therapeutics. The application of the technology is not limited to biopharmaceutical preparations, and, in fact, would benefit any current application that uses HPLC methods for larger biological molecule analyses. The separations technology described will directly lead to useful products for which there is a significant technical and market demand.