Integrated multidimensional chromatography on preparative scale for oligonucleotides purification

IF 3.8 2区 化学 Q1 BIOCHEMICAL RESEARCH METHODS Journal of Chromatography A Pub Date : 2024-10-15 DOI:10.1016/j.chroma.2024.465440
Chiara Nosengo , Desiree Bozza , Giulio Lievore , Sebastian Vogg , Martina Catani , Alberto Cavazzini , Thomas Müller-Späth , Chiara De Luca , Simona Felletti
{"title":"Integrated multidimensional chromatography on preparative scale for oligonucleotides purification","authors":"Chiara Nosengo ,&nbsp;Desiree Bozza ,&nbsp;Giulio Lievore ,&nbsp;Sebastian Vogg ,&nbsp;Martina Catani ,&nbsp;Alberto Cavazzini ,&nbsp;Thomas Müller-Späth ,&nbsp;Chiara De Luca ,&nbsp;Simona Felletti","doi":"10.1016/j.chroma.2024.465440","DOIUrl":null,"url":null,"abstract":"<div><div>Therapeutic oligonucleotides represent a recent breakthrough in the pharmaceutical industry due to their ability to regulate gene expression with great specificity. This aspect allows treatment of a wide range of diseases. However, since oligonucleotides are used for therapeutic purposes, the Active Pharmaceutical Ingredient (API) must fulfill strict purity levels which require intensive purification steps. For oligonucleotides, and biomolecules in general, preparative liquid chromatography is the technique of choice to perform large scale purifications, typically in batch mode, i.e. using a single column. Specifically, since ONs are mainly large, hydrophilic and charged molecules, Anion Exchange chromatography (AEX) and Ion Pair Reversed Phase chromatography (IP-RP) are the preferred chromatographic modes for their downstream processing. Nevertheless, these approaches suffer from a purity-yield trade-off, and for this reason, more than one purification step is usually required. The two chromatographic modes can therefore be used consequently to remove different groups of impurities, thanks to their orthogonality.</div><div>In this work, a multidimensional and orthogonal approach on a (semi)preparative scale, namely “Integrated Batch process”, was applied for the purification of a single-stranded DNA oligonucleotide. This process combines two chromatographic steps without any hold step, operator intervention or sampling of the first step. The performance parameters of the Integrated Batch were compared to those obtained in the single batch runs under different experimental conditions (chromatographic mode, eluent systems), showing the potential of this integrated approach. This proof-of-concept study illustrates how this technique can considerably reduce overall production time and how it allows to increase the robustness and reproducibility of the method, since the process is highly automated.</div></div>","PeriodicalId":347,"journal":{"name":"Journal of Chromatography A","volume":"1737 ","pages":"Article 465440"},"PeriodicalIF":3.8000,"publicationDate":"2024-10-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Chromatography A","FirstCategoryId":"1","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0021967324008148","RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0

Abstract

Therapeutic oligonucleotides represent a recent breakthrough in the pharmaceutical industry due to their ability to regulate gene expression with great specificity. This aspect allows treatment of a wide range of diseases. However, since oligonucleotides are used for therapeutic purposes, the Active Pharmaceutical Ingredient (API) must fulfill strict purity levels which require intensive purification steps. For oligonucleotides, and biomolecules in general, preparative liquid chromatography is the technique of choice to perform large scale purifications, typically in batch mode, i.e. using a single column. Specifically, since ONs are mainly large, hydrophilic and charged molecules, Anion Exchange chromatography (AEX) and Ion Pair Reversed Phase chromatography (IP-RP) are the preferred chromatographic modes for their downstream processing. Nevertheless, these approaches suffer from a purity-yield trade-off, and for this reason, more than one purification step is usually required. The two chromatographic modes can therefore be used consequently to remove different groups of impurities, thanks to their orthogonality.
In this work, a multidimensional and orthogonal approach on a (semi)preparative scale, namely “Integrated Batch process”, was applied for the purification of a single-stranded DNA oligonucleotide. This process combines two chromatographic steps without any hold step, operator intervention or sampling of the first step. The performance parameters of the Integrated Batch were compared to those obtained in the single batch runs under different experimental conditions (chromatographic mode, eluent systems), showing the potential of this integrated approach. This proof-of-concept study illustrates how this technique can considerably reduce overall production time and how it allows to increase the robustness and reproducibility of the method, since the process is highly automated.

Abstract Image

查看原文
分享 分享
微信好友 朋友圈 QQ好友 复制链接
本刊更多论文
制备规模的集成多维色谱法用于寡核苷酸纯化
治疗性寡核苷酸具有高度特异性的基因表达调控能力,是制药业的最新突破。在这方面,寡核苷酸可以治疗多种疾病。然而,由于寡核苷酸用于治疗目的,活性药物成分(API)必须满足严格的纯度要求,这就需要密集的纯化步骤。对于寡核苷酸和一般生物大分子而言,制备型液相色谱法是进行大规模纯化的首选技术,通常采用批处理模式,即使用单一色谱柱。具体来说,由于 ONs 主要是大分子、亲水性分子和带电分子,因此阴离子交换色谱(AEX)和离子对反相色谱(IP-RP)是其下游处理的首选色谱模式。然而,这些方法在纯度-产量之间存在权衡,因此通常需要一个以上的纯化步骤。在这项工作中,我们采用了一种(半)制备规模的多维正交方法,即 "集成批次工艺",用于纯化单链 DNA 寡核苷酸。该工艺将两个色谱步骤结合在一起,第一步没有任何保持步骤、操作员干预或取样。在不同的实验条件(色谱模式、洗脱液系统)下,集成批次的性能参数与单批次运行的性能参数进行了比较,显示了这种集成方法的潜力。这项概念验证研究说明了该技术如何大大缩短整体生产时间,以及如何通过高度自动化的流程提高方法的稳健性和可重复性。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 去求助
来源期刊
Journal of Chromatography A
Journal of Chromatography A 化学-分析化学
CiteScore
7.90
自引率
14.60%
发文量
742
审稿时长
45 days
期刊介绍: The Journal of Chromatography A provides a forum for the publication of original research and critical reviews on all aspects of fundamental and applied separation science. The scope of the journal includes chromatography and related techniques, electromigration techniques (e.g. electrophoresis, electrochromatography), hyphenated and other multi-dimensional techniques, sample preparation, and detection methods such as mass spectrometry. Contributions consist mainly of research papers dealing with the theory of separation methods, instrumental developments and analytical and preparative applications of general interest.
期刊最新文献
Multi–class cyanobacterial toxin analysis using hydrophilic interaction liquid chromatography–mass spectrometry In situ growth of hierarchical porous covalent organic framework coating for enhanced solid-phase microextraction of phenolic compounds Understanding the fundamentals of the on-off retention mechanism of oligonucleotides and their application to high throughput analysis Optimizing conditions in online RPLC × SFC for the analysis of complex samples containing neutral compounds: Solving injection issues. Designing boron-doped carbon dot-functionalized COFs for fluorescence screening and liquid chromatography tandem mass spectrometry detection of toxins
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
现在去查看 取消
×
提示
确定
0
微信
客服QQ
Book学术公众号 扫码关注我们
反馈
×
意见反馈
请填写您的意见或建议
请填写您的手机或邮箱
已复制链接
已复制链接
快去分享给好友吧!
我知道了
×
扫码分享
扫码分享
Book学术官方微信
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术
文献互助 智能选刊 最新文献 互助须知 联系我们:info@booksci.cn
Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。
Copyright © 2023 Book学术 All rights reserved.
ghs 京公网安备 11010802042870号 京ICP备2023020795号-1