Sodium Dodecyl Sulfate Removal Interface to Enable Characterization of Fragment Impurities in Monoclonal Antibodies by Capillary Electrophoresis Sodium Dodecyl Sulfate Coupled to Mass Spectrometry

十二烷基硫酸钠去除接口可通过十二烷基硫酸钠毛细管电泳与质谱联用对单克隆抗体中的片段杂质进行表征

• 批准号: 10759354
• 负责人: Oluwatosin Dada
• 金额: $ 35.89万
• 依托单位: GMJ TECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10759354
• 关键词: Address; Angiotensin I; Angiotensins; Back; Binding; Biological Products; Biotechnology; Blood capillaries; Bovine Serum Albumin; Buffers; Capillary Electrophoresis; Charge; Collaborations; Computer Models; Cost Savings; Coupled; Coupling; Detection; Development; Devices; Digestion; Electrodes; Electrophoresis; Evaluation; Excision; Exclusion; Formulation; Fractionation; Funding; Gel; Glass; Goals; Gold; Human; Immunoglobulin Fragments; Immunoglobulins; Incubated; Industry Standard; Light; Liquid Chromatography; Mainstreaming; Manufacturer; Mass Spectrum Analysis; Modeling; Molecular Sieve Chromatography; Molecular Weight; Monitor; Monoclonal Antibodies; Muramidase; Myoglobin; Noise; Outcome; Ovalbumin; Pattern; Peptide Mapping; Performance; Phase; Polyacrylamide Gel Electrophoresis; Procedures; Process; Production; Proteins; Proteomics; Quality Control; Refractory; Research; Research Personnel; Resolution; Safety; Sampling; Signal Transduction; Silicon; Small Business Innovation Research Grant; Sodium Dodecyl Sulfate; Spectrometry, Mass, Electrospray Ionization; Stress; Structure; Surface; System; Technology; Testing; Universities; Vertebral column; Washington; analog; analytical tool; biopharmaceutical industry; carbonate dehydratase; commercialization; detection limit; disulfide bond reduction; experimental study; fabrication; improved; innovation; instrument; interest; ionization; lithography; manufacture; meetings; migration; nanofabrication; new technology; phase 1 study; programs; protein complex; research and development; solute; therapeutic development; therapeutic protein

PROGRAM SUMMARY Monoclonal antibodies (mAbs) have become mainstream therapeutic proteins in the biopharmaceutical industry (biopharma), and the demand for efficient analytical tools for their characterization and quality control continues to increase. During manufacturing and storage, cleavage of mAb’s primary structure can occur due to different fragmentation induced mechanisms, and the resulting fragment species can have negative implication for the safety and efficacy of the product. Therefore, fragmentation is a critical quality attribute routinely monitored to assess the purity and integrity of mAbs and mAb-derived biologics from production to commercialization. Capillary electrophoresis with sodium dodecyl sulfate (CE-SDS), which is the capillary analogue of polyacrylamide gel electrophoresis (SDS-PAGE), is widely employed for monitoring mAb fragments during production, formulation, stability, and commercial release. CE-SDS employs SDS to denature proteins and render non-covalently associated subunits separable. The separation is based on electrophoretic migration driven by the surface charge induced by SDS binding, which is proportional to the protein’s molecular weight. In a sieving gel matrix, a size-based separation is achieved since all SDS- protein complexes have similar charge-to-size ratio. CE-SDS provides excellent resolution of fragments, but lack of mass identification through direct coupling to mass spectrometry of the resolved fragments currently creates a major gap in the biopharma. GMJ Technologies (GMJ) is requesting SBIR Phase I funding to develop a micro SDS depletion electrophoresis (SDE) interface to allow direct coupling of CE-SDS to mass spectrometry (MS). The proposed interface will use free-flow zone electrophoresis mechanism to remove SDS in-line from the CE-SDS peaks of interest prior to electrospray ionization mass spectrometry (ESI-MS). With this innovation, GMJ aims to address a critical need in the characterization and quality control of mAbs from research to commercialization. By eliminating several laborious steps, the proposed device will significantly improve the efficiency in the characterization workflow for mAbs’ fragments, improve the analysis precision and sensitivity, and provide cost-saving benefits for biopharma researchers.

计划概要 单克隆抗体（mAb）已成为主流治疗蛋白 生物制药行业（生物制药）及其对高效分析工具的需求 表征和质量控制不断增强。在生产和储存过程中， mAb 的一级结构可能由于不同的碎片诱导机制而发生，并且由此产生的 碎片种类可能会对产品的安全性和功效产生负面影响。所以， 碎片是一个关键的质量属性，需要定期监测以评估单克隆抗体的纯度和完整性 以及单克隆抗体衍生的生物制品从生产到商业化的整个过程。 十二烷基硫酸钠毛细管电泳（CE-SDS），即毛细管 聚丙烯酰胺凝胶电泳 (SDS-PAGE) 的类似物，广泛用于监测 mAb 生产、配制、稳定性和商业发布过程中的碎片。 CE-SDS 采用 SDS 来 使蛋白质变性并使非共价结合的亚基可分离。分离是基于 由 SDS 结合诱导的表面电荷驱动的电泳迁移，与 蛋白质的分子量。在筛分凝胶基质中，可以实现基于尺寸的分离，因为所有 SDS- 蛋白质复合物具有相似的电荷大小比。 CE-SDS 提供出色的片段分辨率， 但缺乏通过直接耦合解析片段的质谱进行质量鉴定 目前，生物制药领域存在重大差距。 GMJ Technologies (GMJ) 正在请求 SBIR 第一阶段资金来开发微型 SDS 耗尽电泳 (SDE) 接口，允许 CE-SDS 与质谱 (MS) 直接耦合。 所提出的接口将使用自由流动区带电泳机制从电泳槽中在线去除 SDS 在电喷雾电离质谱 (ESI-MS) 之前获得感兴趣的 CE-SDS 峰。凭借这一创新， GMJ 旨在满足单克隆抗体从研究到生产过程中表征和质量控制的关键需求。 商业化。通过消除几个费力的步骤，所提出的设备将显着改善 mAb 片段表征工作流程的效率，提高分析精度 灵敏度，并为生物制药研究人员提供节省成本的好处。