Plasma proteomic profiling identifies apolipoprotein A4 as a downregulated biomarker of adrenocortical carcinoma: a multi-platform discovery and validation study.

OBJECTIVES

Adrenocortical carcinoma (ACC) is a rare, aggressive malignancy associated with heterogeneous prognosis. Preoperative differentiation from adrenocortical adenoma (ACA) remains challenging, and no serum tumor marker has been established. We aimed to identify circulating protein biomarkers that distinguish ACC from ACA using a stepwise, multiplatform proteomics strategy.

METHODS

We assembled discovery (ACC = 10, ACA = 67) and verification (ACC = 7, ACA = 11) cohorts from a tertiary center and profiled fasting plasma using liquid chromatography-mass spectrometry (LC-MS/MS) with data-independent acquisition. Differentially expressed proteins (DEPs) were defined by t-tests with P < .05 and |fold-change| >1.2; DEPs common to both cohorts were prioritized. Targeted validation by parallel reaction monitoring (PRM) used an expanded, two-center cohort including additional cases from Asan Medical Center (ACC = 31; ACA = 78). Orthogonal validation employed the Olink Explore 384 Inflammation II panel in an independent set (ACC = 15; ACA = 24).

RESULTS

The discovery cohort yielded 67 DEPs (22 upregulated and 45 downregulated in ACC), and the verification cohort identified 17 DEPs. Three proteins, CD44, proteoglycan 4, and apolipoprotein A4 (APOA4), were common to both analyses and were underexpressed in ACC compared with ACA. In PRM, CD44 and APOA4 showed directionally concordant, significant decreases in ACC, prioritizing these markers for further evaluation. In the Olink analysis, 40 proteins differed between ACC and ACA after false discovery rate correction; APOA4 remained significantly lower in ACC.

CONCLUSION

Across discovery, targeted, and orthogonal platforms, APOA4 consistently exhibited lower circulating levels in ACC, supporting its potential as a serum biomarker for the preoperative differentiation of ACC from ACA. External, multiethnic validation and clinically deployable assays, alone or within multimarker panels, are warranted.