Center for Translational Medicine and Pharmacology, Icahn School of Medicine at Mount Sinai, New York, United States.
Center for Biomedical Research, Population Council, New York, United States.
Elife. 2023 Jun 19;12:e88898. doi: 10.7554/eLife.88898.
Highly concentrated antibody formulations are oftentimes required for subcutaneous, self-administered biologics. Here, we report the development of a unique formulation for our first-in-class FSH-blocking humanized antibody, MS-Hu6, which we propose to move to the clinic for osteoporosis, obesity, and Alzheimer's disease. The studies were carried out using our Good Laboratory Practice (GLP) platform, compliant with the Code of Federal Regulations (Title 21, Part 58). We first used protein thermal shift, size exclusion chromatography, and dynamic light scattering to examine MS-Hu6 concentrations between 1 and 100 mg/mL. We found that thermal, monomeric, and colloidal stability of formulated MS-Hu6 was maintained at a concentration of 100 mg/mL. The addition of the antioxidant L-methionine and chelating agent disodium EDTA improved the formulation's long-term colloidal and thermal stability. Thermal stability was further confirmed by Nano differential scanning calorimetry (DSC). Physiochemical properties of formulated MS-Hu6, including viscosity, turbidity, and clarity, confirmed with acceptable industry standards. That the structural integrity of MS-Hu6 in formulation was maintained was proven through Circular Dichroism (CD) and Fourier Transform Infrared (FTIR) Spectroscopy. Three rapid freeze-thaw cycles at -80 °C/25 °C or -80 °C/37 °C further revealed excellent thermal and colloidal stability. Furthermore, formulated MS-Hu6, particularly its Fab domain, displayed thermal and monomeric storage stability for more than 90 days at 4°C and 25°C. Finally, the unfolding temperature (T) for formulated MS-Hu6 increased by >4.80 °C upon binding to recombinant FSH, indicating highly specific ligand binding. Overall, we document the feasibility of developing a stable, manufacturable and transportable MS-Hu6 formulation at a ultra-high concentration at industry standards. The study should become a resource for developing biologic formulations in academic medical centers.
高浓度的抗体制剂通常用于皮下、自我给药的生物制剂。在这里，我们报告了我们的首个 FSH 阻断人源化抗体 MS-Hu6 的独特制剂的开发，我们提议将其推向骨质疏松症、肥胖症和阿尔茨海默病的临床应用。这些研究是在符合联邦法规（标题 21，第 58 部分）的我们的良好实验室规范（GLP）平台上进行的。我们首先使用蛋白质热移位、尺寸排阻色谱和动态光散射来检查 1 至 100mg/ml 之间的 MS-Hu6 浓度。我们发现，在 100mg/ml 的浓度下，配制的 MS-Hu6 的热、单体和胶体稳定性得以维持。抗氧化剂 L-蛋氨酸和螯合剂 disodium EDTA 的添加改善了制剂的长期胶体和热稳定性。通过纳米差示扫描量热法（DSC）进一步证实了热稳定性。配制的 MS-Hu6 的物理化学性质，包括粘度、浊度和清晰度，均符合可接受的行业标准。通过圆二色性（CD）和傅里叶变换红外（FTIR）光谱证明，MS-Hu6 的结构完整性在制剂中得以保持。经过三次-80°C/25°C 或-80°C/37°C 的快速冻融循环进一步显示出优异的热和胶体稳定性。此外，配制的 MS-Hu6，特别是其 Fab 结构域，在 4°C 和 25°C 下储存 90 天以上，仍保持热和单体储存稳定性。最后，在与重组 FSH 结合后，配制的 MS-Hu6 的展开温度（T）升高了>4.80°C，表明具有高度特异性的配体结合。总之，我们证明了在行业标准下开发高浓度稳定、可制造和可运输的 MS-Hu6 制剂的可行性。该研究应成为学术医学中心开发生物制剂制剂的资源。
