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Development and Application of a Liquid Chromatography-Mass Spectrometry Method for Residual Iodixanol Quantification in AAV-Based Gene Therapy Product Development
Human Gene Therapy ( IF 4.2 ) Pub Date : 2022-01-17 , DOI: 10.1089/hum.2021.136
Yi Pu 1 , Russell Katz 2 , Yunqiu Chen 1 , Vic Kostrubsky 3 , Pete Clarner 4 , Shih-Ching Lo 5 , Zoran Sosic 1 , Bernice Yeung 1
Affiliation  

Adeno-associated viruses (AAVs) are nonenveloped viruses that have become popular gene transfer vectors to deliver DNA to target cells in clinical gene therapy. Iodixanol-based density gradient is one of the widely used purification methods for serotype-independent AAVs. However, residual iodixanol in AAV could be a safety concern, and further purification to remove this process-related impurity is typically needed. An analytical assay with high sensitivity is essential for the detection of residual iodixanol to ensure the safety of AAV products. We developed a liquid chromatography-mass spectrometry method with the limit of quantification of 0.01 μg/mL for residual iodixanol measurement in AAVs. The method also demonstrated linearity over four orders of magnitude that allows quantifying a high iodixanol concentration in in-process samples with excellent recovery and accuracy. In addition, we further explored a highly efficient purification method for removal of the residual iodixanol, to minimize the safety concern from iodixanol as a process impurity.

中文翻译:

基于 AAV 的基因治疗产品开发中残留碘克沙醇定量液相色谱-质谱法的开发和应用

腺相关病毒 (AAV) 是无包膜病毒,已成为流行的基因转移载体,可在临床基因治疗中将 DNA 递送至靶细胞。基于碘克沙醇的密度梯度是血清型非依赖性 AAV 广泛使用的纯化方法之一。然而,AAV 中残留的碘克沙醇可能是一个安全问题,通常需要进一步纯化以去除这种与工艺相关的杂质。具有高灵敏度的分析测定对于检测残留碘克沙醇以确保 AAV 产品的安全性至关重要。我们开发了一种液相色谱-质谱法,定量限为 0.01 μg/mL,用于 AAV 中残留的碘克沙醇测量。该方法还证明了超过四个数量级的线性,允许以出色的回收率和准确性量化过程中样品中的高浓度碘克沙醇。此外,我们进一步探索了一种去除残留碘克沙醇的高效纯化方法,以最大限度地减少碘克沙醇作为工艺杂质带来的安全隐患。
更新日期:2022-01-20
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