Poster - #UCLA-100 - Keywords: Hepatocellular carcinoma; extracellular vesicle; biomarker; early detection; surface protein.

An Extracellular Vesicle Surface Protein Assay for Early Detection of Hepatocellular Carcinoma

Yi-Te Lee, MD; Na Sun, PhD; Ceng Zhang, MD; Ryan Y. Zhang, BS; Benjamin V. Tran, MD; Jasmine J Wang, MD; Sungyong You, PhD; *Ju Dong Yang, MD, MS; Vatche G Agopian, MD; Hsian-Rong Tseng, PhD; and Yazhen Zhu, MD, PhD.
1Department of Molecular and Molecular Pharmacology, 2Department of Surgery, David Geffen School of Medicine, 5Jonsson Comprehensive Cancer Center, University of California, Los Angeles, CA; 3Division of Cancer Systems Biology, Department of Surgery, 4Karsh Division of Gastroenterology and Hepatology, Cedars Sinai Medical Center, Los Angeles, CA.

ABSTRACT:
Background: The sensitivity of current surveillance methods for detection of early-stage hepatocellular carcinoma (HCC) is suboptimal. Extracellular vesicles (EVs) are nanoparticles containing biomolecules such as DNA, RNA, and proteins. Given their presence in circulation during tumorigenesis, profiling tumor-derived EVs is regarded as a promising liquid biopsy strategy for early cancer detection. In this study, we aim to develop an HCC EV-based surface protein assay for detecting early-stage HCC. Methods: The expression of four potential HCC-associated surface protein markers, EpCAM, ASGPR1, CD147, and GPC3, was first evaluated using a 708-case HCC tissue microarray and EVs derived from HepG2 and Hep3B cells. An HCC EV Surface Protein Assay, composed of covalent chemistry-mediated HCC EV purification and real-time immuno-PCR analysis, was then developed and optimized for quantifying subpopulations of HCC EVs by adopting the four surface protein markers. Utilizing logistic regression, an HCC EV ECG was established for differentiating early-stage HCC from liver cirrhosis in a training cohort (n=106), and validated in an independent external cohort (n=72). Results: Overall, 99.7% of tissue microarray stained positive for at least one of the four HCC-associated protein markers, which were subsequently validated in HCC EVs. In the training cohort, HCC EV ECG score demonstrated an area under the receiver operating curve (AUROC) of 0.95 (95% CI=0.90-0.99) for distinguishing early-stage HCC from cirrhosis with a sensitivity of 91% and a specificity of 90%. The AUROCs of HCC EV ECG score remained excellent in the validation cohort (0.93, 95% CI=0.87-0.99), and the subgroups by etiology (viral: 0.95, 95% CI=0.90-1.00; nonviral: 0.94, 95% CI=0.88-0.99). Conclusions: An HCC EV ECG score based on immuno-PCR based analysis of HCC EV surface proteins were developed and validated for accurately detecting early-stage HCC from cirrhosis. HCC EV ECG score holds great promise to augment current surveillance method to identify more patients with HCC at a curable stage and improve their long-term outcomes.