Northeast Medical Products Laboratory, Office of Regulatory Science, Office of Regulatory Affairs, U.S. Food and Drug Administration, Jamaica, NY 11433, USA.
Division of Complex Drug Analysis, Office of Testing and Research, Office of Pharmaceutical Quality, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, MD 20993, USA.
Molecules. 2021 Jul 13;26(14):4251. doi: 10.3390/molecules26144251.
Peptide and protein drug molecules fold into higher order structures (HOS) in formulation and these folded structures are often critical for drug efficacy and safety. Generic or biosimilar drug products (DPs) need to show similar HOS to the reference product. The solution NMR spectroscopy is a non-invasive, chemically and structurally specific analytical method that is ideal for characterizing protein therapeutics in formulation. However, only limited NMR studies have been performed directly on marketed DPs and questions remain on how to quantitively define similarity. Here, NMR spectra were collected on marketed peptide and protein DPs, including calcitonin-salmon, liraglutide, teriparatide, exenatide, insulin glargine and rituximab. The 1D H spectral pattern readily revealed protein HOS heterogeneity, exchange and oligomerization in the different formulations. Principal component analysis (PCA) applied to two rituximab DPs showed consistent results with the previously demonstrated similarity metrics of Mahalanobis distance (D) of 3.3. The 2D H-C HSQC spectral comparison of insulin glargine DPs provided similarity metrics for chemical shift difference (Δδ) and methyl peak profile, i.e., 4 ppb for H, 15 ppb for C and 98% peaks with equivalent peak height. Finally, 2D H-N sofast HMQC was demonstrated as a sensitive method for comparison of small protein HOS. The application of NMR procedures and chemometric analysis on therapeutic proteins offer quantitative similarity assessments of DPs with practically achievable similarity metrics.
肽和蛋白质药物分子在制剂中折叠成高级结构（HOS），这些折叠结构通常对药物的疗效和安全性至关重要。仿制药或生物类似药产品（DPs）需要表现出与参比产品相似的 HOS。溶液 NMR 光谱是一种非侵入性、化学和结构特异性的分析方法，非常适合于制剂中蛋白质治疗剂的特征描述。然而，只有有限的 NMR 研究直接针对市售 DPs 进行，如何定量定义相似性仍存在疑问。在这里，对市售的肽和蛋白质 DPs 进行了 NMR 光谱收集，包括鲑鱼降钙素、利拉鲁肽、特立帕肽、艾塞那肽、甘精胰岛素和利妥昔单抗。一维 H 谱图模式容易揭示不同制剂中蛋白质 HOS 的异质性、交换和寡聚化。主成分分析（PCA）应用于两种利妥昔单抗 DPs 显示出与先前证明的相似性度量——马哈拉诺比斯距离（D）为 3.3 的一致结果。甘精胰岛素 DPs 的二维 H-C HSQC 光谱比较提供了化学位移差（Δδ）和甲基峰轮廓的相似性度量，即 H 为 4 ppb、C 为 15 ppb 和 98%的峰具有等效的峰高。最后，二维 H-N sofast HMQC 被证明是一种用于比较小蛋白质 HOS 的敏感方法。NMR 程序和化学计量分析在治疗性蛋白质中的应用为 DPs 提供了定量相似性评估，具有实际可行的相似性度量。
