Urinary Proteomics Identifies ADAMDEC1 as a Non-invasive Biomarker of Unstable Carotid Atherosclerotic Plaques.

Abstract

Rupture of vulnerable carotid atherosclerotic plaques (CAPs) is a major precipitating cause of ischemic stroke. Current imaging-based risk stratification is useful for anatomical and structural assessment, but it does not readily provide noninvasive access to plaque-associated molecular information. Here, we aimed to identify and validate a noninvasive urinary biomarker of CAP instability and to explore its biological relevance. We employed a multi-stage study design integrating untargeted urinary proteomics data-independent acquisition mass spectrometry in a discovery cohort of 179 patients with systematic interrogation of plaque tissue proteomic and transcriptomic datasets. Candidate proteins were further evaluated by ELISA in an independent cohort of 225 individuals, including patients with stable or unstable CAPs and healthy controls. Cellular and tissue-level analyses included single-cell RNA sequencing re-analysis, human plaque validation by immunohistochemistry, western blotting, and immunofluorescence, as well as in vitro macrophage phenotypic assays. Urinary proteomic profiling identified 227 shared differential proteins associated with plaque instability. In the independent validation cohort, urinary ADAM-Like Decysin 1 (ADAMDEC1) was confirmed as a candidate urinary marker, showing a stepwise increase from healthy controls to stable and unstable CAPs. Urinary ADAMDEC1 showed robust discriminatory performance for unstable plaques (AUC = 0.882, 95% CI 0.822-0.941). Higher urinary ADAMDEC1 levels were associated with symptomatic status and plaque vulnerability features, while higher intraplaque ADAMDEC1 expression was associated with adverse cerebrovascular outcomes. Single-cell and human plaque analyses showed that ADAMDEC1 was enriched in inflammatory macrophage-associated plaque regions, and in vitro assays supported an association between ADAMDEC1 and pro-inflammatory macrophage polarization. Urinary ADAMDEC1 is a noninvasive biomarker associated with carotid plaque instability. Integrated tissue, single-cell, and macrophage assays support a link between ADAMDEC1 and macrophage-associated inflammatory features in vulnerable plaques.