PO.CH02.01 · 化学
推进血浆蛋白质组学：一种用于结直肠癌深度定量生物标志物发现的新一代纳米颗粒富集工作流程
Advancing plasma proteomics: A next-generation nanoparticle enrichment workflow for deep and quantitative biomarker discovery in colorectal cancer
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摘要 Abstract
中文摘要
人血浆是一种微创的循环癌症生物标志物来源，但其极大的蛋白动态范围和个体间异质性阻碍了对低丰度肿瘤来源标志物的检测。在结直肠癌（CRC）中，早期检测、风险分层和治疗监测仍是重大的未满足需求，这些分析上的障碍限制了临床转化。在此，我们提出一套单颗粒富集工作流程，将P2纳米颗粒与iST样本制备和Spectronaut® 20 DIA分析相结合，以在人EDTA血浆生物标志物发现中实现更深的蛋白质组覆盖、更高的定量精度和更强的生物学洞见。来自CRC和健康标本的人EDTA血浆样本（每组n = 6）经一项初步研究，采用P2-iST血浆工作流程（PreOmics）进行处理，该工作流程采用专有纳米颗粒，可选择性捕获低丰度蛋白同时去除高丰度蛋白。随后使用优化的iST技术（PreOmics）对富集样本进行消化和纯化，确保标准化的肽段生成。肽段在以dia-PASEF®模式运行的timsTOF HT（Bruker）上进行分析，数据使用Spectronaut® 20（Biognosys）通过directDIA+框架进行处理，以实现深度定量分析。与未富集的血浆样本相比，该整合工作流程产生了更深的蛋白质组覆盖和高定量精度（重复样本间蛋白水平中位CV<15%）。主成分分析和层次聚类分析显示CRC与对照样本之间具有清晰的区分。功能富集分析揭示了区分疾病组与对照组的不同生物学通路，凸显了该工作流程的灵敏度和判别潜力。P2-iST血浆工作流程整合了基于纳米颗粒的富集、标准化消化和先进的DIA分析，以生成深度、精确且可重复的血浆蛋白质组，反映结直肠癌中与疾病相关的生物学。通过开启循环生物标志物发现的新可能，它支持早期检测、分子分型、预后和治疗监测方面的研究。其可扩展、易于自动化的设计进一步支持在结直肠癌及其他癌症中开展大队列验证和纵向研究。
查看英文原文 English abstract
Human plasma provides a minimally invasive source of circulating cancer biomarkers, yet its extreme protein dynamic range and interindividual heterogeneity hinder the detection of low-abundance, tumor-derived markers. In colorectal cancer (CRC), where early detection, risk stratification, and therapy monitoring remain major unmet needs, these analytical barriers limit clinical translation. Here, we present a single-particle enrichment workflow that combines P2 nanoparticles with iST sample preparation and Spectronaut® 20 DIA analysis to enable deeper proteome coverage, improved quantitative precision, and enhanced biological insights in human EDTA-plasma biomarker discovery. Human EDTA-plasma samples from a pilot study of CRC and healthy specimens (n = 6 per group) were processed using the P2-iST Plasma workflow (PreOmics), employing proprietary nanoparticles that selectively capture low-abundance proteins while depleting high-abundance species. Enriched samples were subsequently digested and purified using the optimized iST technology (PreOmics), ensuring standardized peptide generation. Peptides were analyzed on a timsTOF HT (Bruker) operated in dia-PASEF® mode and data processed in Spectronaut® 20 (Biognosys) using the directDIA+ framework for deep, quantitative profiling. Compared with non-enriched plasma samples, the integrated workflow yielded deeper proteome coverage and high quantitative precision (median protein-level CV <15% across replicates). Principal-component and hierarchical clustering analyses demonstrated clear discrimination between CRC and control samples. Functional enrichment revealed distinct biological pathways differentiating disease from control groups, highlighting the sensitivity and discriminative potential of the workflow. The P2-iST Plasma workflow integrates nanoparticle-based enrichment, standardized digestion, and advanced DIA analysis to generate deep, precise, and reproducible plasma proteomes that reflect disease-relevant biology in colorectal cancer. By unlocking new possibilities in circulating biomarker discovery, it supports research in early detection, molecular subtyping, prognosis, and treatment monitoring. Its scalable, automation-friendly design further enables large-cohort validation and longitudinal studies in colorectal and other cancers.
利益披露 Disclosure
C. Ellis, None..
K. Limm, None..
S. Schär, None..
R. Bruderer, None..
N. Kulak, None.