Abstract 7665: Detection of γ-OHPdG as a hepatocellular carcinoma recurrence biomarker by CTCs and LC-MS/MS
Authors: Kuo Mark , Aktop Merve Ozge , Ghabi Elie , Ouf Mahmoud A. , Carder Margaret , Muralidaran Vinona , Ascue Jhalen , Hlouskova Marketa , Wu Pengfei , Cheng Guang , Hecht Stephen S. , Fang Hong-Bin , Hanna Nader , He Jin , Kroemer Alexander H. , Aggarwal Monika , He Aiwu R. , Chung Fung-Lung 
Published: March 2024

Abstract
Background: Hepatocellular carcinoma (HCC) is the third leading cause of cancer death globally. Recurrence risks following curative resection remain high, ~50% after three years and 75-100% over five years. A biomarker for HCC recurrence is urgently needed to help lead treatment strategies. We previously showed that hepatic γ-hydroxy-1, N2-propano-2′-deoxyguanosine (γ-OHPdG), a DNA adduct derived from lipid peroxidation, increased during hepatocarcinogenesis. Furthermore, in a retrospective study, higher γ-OHPdG levels in post-surgical liver tumor tissues were associated with poorer survival (p&lt;0.0001) and shorter recurrence-free survival (p=0.007), suggesting that it may be a prognostic biomarker for HCC (Hepatology 2018). In this study, we aimed to test γ-OHPdG detected in livers after curative resection as a recurrence biomarker in a prospective study and, in addition, we developed a new non-invasive method for γ-OHPdG in circulating tumor cells (CTCs). This multicenter study seeks to validate γ-OHPdG as a biomarker for HCC recurrence.
Methods: A total of 28 HCC patients undergoing hepatectomy at MedStar/Georgetown University Hospital, Johns Hopkins University Hospital, and the University of Maryland Medical Center were enrolled. γ-OHPdG was detected in DNA isolated from hepatic tumors and non-tumors by a previous liquid chromatograph-tandem mass spectrometry (LC-MS/MS) method and in HCC CTCs isolated from blood samples. CTCs were isolated using hepatocyte-specific rabbit anti-asialoglycoprotein-1 (ASGPR-1), followed by staining with mouse anti-γ-OHPdG antibodies. Clinical outcomes were correlated with γ-OHPdG in liver tissues and CTCs.
Results: In our current cohort of 28 patients (mean age: 64.7 years; 82% male), five patients experienced HCC recurrence, with an estimated progression-free survival (PFS) rate of 85% over 8 months. γ-OHPdG levels (adducts/109 nucleotides) detected by LC-MS/MS varied widely in tumor tissues (5-56, mean: 11.8) and non-tumor tissue (7-71, mean: 20.6). Preliminary analysis showed shorter PFS associated with γ-OHPdG in non-tumors (cutoff: 16, p=0.0271). γ-OHPdG (arbitrary units x107) in CTCs also varied widely in the nucleus (0.95-18.84, mean: 2.85) and in the whole cell (0.16- 52.18, mean: 9.72). Total CTC γ-OHPdG intensity in both the nucleus (p=0.006) and in the whole cell (p=0.005) was associated with its levels in non-tumor tissue. Furthermore, a shorter PFS was associated with CTC count (cutoff: 270 cells, p=0.01), and total CTC whole cell γ-OHPdG intensity (cutoff: 21.17 × 107 arbitrary units, p=0.002).
Conclusion: Preliminary results suggest that γ-OHPdG, measured in tissues by LC-MS/MS and in blood by CTC, predicts HCC recurrence. Once evaluated in all enrolled subjects, multiple variant analyses will be carried out to include stage, AFP, grade of differentiation, vascular invasion on surgical samples, and status of viral hepatitis infection.
Citation Format: Mark Kuo, Merve Ozge Aktop, Elie Ghabi, Mahmoud A. Ouf, Margaret Carder, Vinona Muralidaran, Jhalen Ascue, Marketa Hlouskova, Pengfei Wu, Guang Cheng, Stephen S. Hecht, Hong-Bin Fang, Nader Hanna, Jin He, Alexander H. Kroemer, Monika Aggarwal, Aiwu R. He, Fung-Lung Chung. Detection of γ-OHPdG as a hepatocellular carcinoma recurrence biomarker by CTCs and LC-MS/MS [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7665.