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采用自上而下的差示质谱和代谢物数据库匹配法鉴定治疗性肽和蛋白质的代谢物。

Metabolite Identification of Therapeutic Peptides and Proteins by Top-down Differential Mass Spectrometry and Metabolite Database Matching.

出版信息

Anal Chem. 2020 Jun 16;92(12):8298-8305. doi: 10.1021/acs.analchem.0c00652. Epub 2020 May 28.

DOI:10.1021/acs.analchem.0c00652
PMID:32402188
Abstract

As metabolism impacts the efficacy and safety of therapeutic peptides and proteins (TPPs), understanding of the metabolic fate of TPPs is critical for their preclinical and clinical development. Despite the continued increase of new TPPs entering clinical trials, the metabolite identification (MetID) of these emerging modalities remains challenging. In the present study, we report an analytical workflow for MetID of TPPs. Using insulin detemir as an example, we demonstrated that top-down differential mass spectrometry (dMS) was able to distinguish and discover metabolites from complex biological matrices. For structural interpretation, we developed an algorithm to generate a complete and nonredundant theoretical metabolite database for a TPP of any topology (e.g., branched, multicyclic, etc.). Candidate structures of a metabolite were obtained by matching the monoisotopic mass of a dMS feature to the theoretical metabolite database. Finally, the MS/MS sequence tags enabled unambiguous characterization of metabolite structures when isobaric/isomeric candidates were present. This platform is widely applicable to TPPs with complex structures and will ultimately guide the structural optimization of TPPs in pharmaceutical development.

摘要

由于代谢会影响治疗性肽和蛋白质（TPPs）的疗效和安全性，因此了解 TPPs 的代谢命运对于它们的临床前和临床开发至关重要。尽管越来越多的新 TPP 进入临床试验，但这些新兴模式的代谢产物鉴定（MetID）仍然具有挑战性。在本研究中，我们报告了一种用于 TPP 的 MetID 的分析工作流程。我们以胰岛素地特为例，证明了自上而下的差示质谱（dMS）能够区分和发现复杂生物基质中的代谢产物。为了进行结构解释，我们开发了一种算法，用于为任何拓扑结构的 TPP（例如分支、多环等）生成完整且无冗余的理论代谢物数据库。通过将 dMS 特征的单同位素质量与理论代谢物数据库进行匹配，可以获得代谢物的候选结构。最后，当存在等压/等构象候选物时，MS/MS 序列标签可用于明确表征代谢物结构。该平台广泛适用于具有复杂结构的 TPP，并将最终指导药物开发中 TPP 的结构优化。

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