Protein Aggregation in Amorphous Solids

描述(由申请人提供)：蛋白质药物是制药行业增长最快的领域之一。对这些药物的强劲需求反映出它们有能力治疗以前难以治疗的疾病，包括癌症、传染病、自身免疫性疾病和心血管疾病。目前市场上40%以上的蛋白质药物产品是无定形固体，这种形式通常是为了延长货架期和保持效力而选择的。然而，蛋白质药物在固体状态下经历了各种物理和化学降解过程。聚合是这些过程中最常见的过程之一。由于聚集体的存在与效力的降低和威胁生命的免疫原性副作用的可能性增加有关，因此必须在制造和储存过程中检测并移除它们。这增加了生产蛋白质药物的成本，最终增加了患者的成本，并排除了无法有效稳定的有前景的新蛋白质药物的商业化。该研究计划的目标是基于对所涉及的化学(即共价)和物理(即非共价)机制的透彻理解，开发合理的方法来防止蛋白质在固态中聚集。核心假设是蛋白质在无定形固体中的聚集是特定的共价反应和/或蛋白质序列中容易聚集的热点的暴露的结果，这两者都可以通过设计固体环境来防止。为具体目标1提出的研究将阐明无定形固体中硫醇-二硫化物交换和二硫化物扰乱的机制，并将确定控制这些反应的固体性质。这些研究验证了一种假设，即这些常见的共价聚集路径在溶液和固体中倾向于不同的路径，并受固体组成的影响。特定目标2将利用氢/氢(H/D)交换和分子动力学模拟(MDS)来确定非共价蛋白质在无定形固体中聚集的“热点”。这项工作测试了一种假设，即无定形固体中这些定量的、高分辨率的蛋白质结构测量将与长期储存期间的非共价聚集相关。特殊目标3将开发一种计算模型，根据蛋白质和固体的性质预测蛋白质在无定形固体中的聚集，产生一种用于配方设计和识别对防止聚集至关重要的变量的工具。这项工作与NIH提高国家保护和改善健康的能力的使命有关，因为它解决了保存快速增长的一类药物的效力和安全性的方法。这项工作也与该机构的目标一致，即通过提供工具和知识，将活性蛋白质开发成可销售的药物产品，以确保公共研究投资继续获得高回报。蛋白质药物产品中聚集体的存在增加了当给患者使用药物时威胁生命的免疫原性反应的可能性。了解无定形固体中聚集体的形成将有助于确保这一快速增长的药物类别的安全性。这项工作还将通过为蛋白质药物配方提供合理的基础，帮助控制药物开发成本。

DESCRIPTION (provided by applicant): Protein drugs are one of the fastest growing segments of the pharmaceutical industry. The strong demand for these drugs reflects their ability to treat previously intractable diseases, including cancers, infectious disease, autoimmune disorders and cardiovascular disease. More than 40% of currently marketed protein drug products are amorphous solids, a form often chosen to prolong shelf-life and preserve potency. Nevertheless, protein drugs undergo a variety of physical and chemical degradation processes in the solid state. Aggregation is one of the most common of these processes. Since the presence of aggregates is associated with decreased potency and with an increased potential for life-threatening immunogenic side effects, they must be detected and removed during manufacturing and storage. This adds to the cost of producing protein drugs, ultimately increasing the cost to the patient and precluding the commercialization of promising new protein drugs that cannot be stabilized effectively. The goal of this research program is to develop rational methods for preventing protein aggregation in the solid state based on a thorough understanding of the chemical (i.e., covalent) and physical (i.e., non- covalent) mechanisms involved. The central hypothesis is that protein aggregation in amorphous solids is the result of specific covalent reactions and/or the exposure of aggregation-prone "hot spots" in the protein sequence, both of which can be prevented by designing the solid environment. Studies proposed for Specific Aim 1 will elucidate the mechanisms of thiol-disulfide exchange and disulfide scrambling in amorphous solids and will identify solid properties that control these reactions. The studies test the hypothesis that these common routes of covalent aggregation favor different pathways in solution and in the solid state and are influenced by solid composition. Specific Aim 2 will identify "hot spots" for non-covalent protein aggregation in amorphous solids using hydrogen/deuterium (H/D) exchange and molecular dynamics simulation (MDS). The work tests the hypothesis that these quantitative, high resolution measures of protein structure in amorphous solids will correlate with non-covalent aggregation during long-term storage. Specific Aim 3 will develop a computational model that predicts protein aggregation in amorphous solids based on properties of the protein and solid, producing a tool for formulation design and identifying variables critical to preventing aggregation. The work is relevant to the NIH mission of advancing the Nation's capacity to protect and improve health in that it addresses methods to preserve the potency and safety of a rapidly growing class of drugs. The work is also consistent with the agency's goal of ensuring a continued high return on the public investment in research by providing tools and knowledge for developing active proteins into marketable drug products. The presence of aggregates in protein drug products increases the potential for life-threatening immunogenic responses when the drugs are administered to patients. Understanding aggregate formation in amorphous solids will help ensure the safety of this rapidly growing drug class. The work will also help to control drug development costs by providing a rational basis for protein drug formulation.