Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related death worldwide. Most cases of HCC develop in patients that already have liver cirrhosis and have been recommended for surveillance for an early onset of HCC. Only 20-30% of patients with HCC are currently eligible for curative therapy with surgical resection where the impact of these therapeutic interventions on the overall survival of patients with HCC is related to the detection of HCC at earlier stages. With the development of new markers one would hope to increase the proportion of patients that can be offered curative therapies and significantly impact the survival of patients with this disease. In our previous work we have found several glycoprotein markers of early stage HCC versus cirrhosis which can provide a performance that is potentially better than that of AFP or together with AFP can improve its overall performance. In particular a bifucosylated glycan structure found in haptoglobin from HCC patients is undergoing a phase 2 EDRN blinded validation where this marker appears to improve the performance for detection of early stage HCC of all etiologies. A new strategy that has been investigated is to screen serum for large numbers of core-fucosylated(CF) sites using mass spectrometry where over 1300 sites have been detected in the serum of HCC patients. We have found several CF-glycopeptides in serum that can distinguish between early HCC and cirrhosis and which are etiology specific for HCV-HCC versus HCV-cirrhosis and ALC-HCC versus ALC-cirrhosis. In the work proposed herein, we will develop a mass spec based assay that is specific for detecting candidate CF- glycopeptides that are differentially expressed in serum of early HCC patients versus cirrhosis. It involves digestion of proteins in total serum to glycopeptides followed by enzymatic removal of the glycan down to the GlcNAc-Fuc core fucosylation. The CF-glycopeptides are labeled with isotopic tags and analyzed by a novel glyco-MRM(multiple reaction monitoring) assay using tandem mass spectrometry where differences in CF expression between HCC and cirrhosis can be detected. This method can be multiplexed for many markers simultaneously, does not require the use of antibodies and can be automated. The hypothesis here is that these structural changes depend on a different mechanism than protein level assays and could provide complementary information to the ELISA assay for AFP where together they may provide an improved AUC. We will test these markers in a confirmation set of 300 samples and evaluate their performance together with the ELISA assay for AFP and a mass spec assay for bifucosylated Hp. The markers that are found to improve on the performance of AFP either alone or in combination with AFP or the bifucosylated Hp assay will then undergo a blinded validation in the phase 2 EDRN validation set. Markers that perform successfully in this blinded validation will then go on for possible use in future work in the HEDS study which is a large scale prospective-retrospective evaluation of markers for early detection of HCC.

Public Health Relevance

If this project is successful then it will result in identification of site-specific core-fucosylated(CF)- glycopeptides from glycosylated proteins in patient serum that can distinguish early stage HCC from cirrhosis as validated in the EDRN phase 2 set. This will be a major advance in the field if these potential markers pass initial validation using the CF-glyco-MRM mass spec based assay to be developed in this work and these markers will potentially be used in future work in a larger EDRN funded study for early detection of HCC.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA160254-09
Application #
9959332
Study Section
Cancer Biomarkers Study Section (CBSS)
Program Officer
Rinaudo, Jo Ann S
Project Start
2012-05-01
Project End
2021-06-30
Budget Start
2020-07-01
Budget End
2021-06-30
Support Year
9
Fiscal Year
2020
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Surgery
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Yin, Haidi; An, Mingrui; So, Pui-Kin et al. (2018) The analysis of alpha-1-antitrypsin glycosylation with direct LC-MS/MS. Electrophoresis 39:2351-2361
Huang, Yifan; Zhou, Shiyue; Zhu, Jianhui et al. (2017) LC-MS/MS isomeric profiling of permethylated N-glycans derived from serum haptoglobin of hepatocellular carcinoma (HCC) and cirrhotic patients. Electrophoresis 38:2160-2167
Yin, Haidi; Zhu, Jianhui; Wu, Jing et al. (2016) A procedure for the analysis of site-specific and structure-specific fucosylation in alpha-1-antitrypsin. Electrophoresis 37:2624-2632
Zhu, Jianhui; Wu, Jing; Yin, Haidi et al. (2015) Mass Spectrometric N-Glycan Analysis of Haptoglobin from Patient Serum Samples Using a 96-Well Plate Format. J Proteome Res 14:4932-9
Yin, Haidi; Tan, Zhijing; Wu, Jing et al. (2015) Mass-Selected Site-Specific Core-Fucosylation of Serum Proteins in Hepatocellular Carcinoma. J Proteome Res 14:4876-84
Zhang, Yiwei; Zhu, Jianhui; Yin, Haidi et al. (2015) ESI-LC-MS Method for Haptoglobin Fucosylation Analysis in Hepatocellular Carcinoma and Liver Cirrhosis. J Proteome Res 14:5388-95
Tan, Zhijing; Yin, Haidi; Nie, Song et al. (2015) Large-scale identification of core-fucosylated glycopeptide sites in pancreatic cancer serum using mass spectrometry. J Proteome Res 14:1968-78
Wu, Jing; Zhu, Jianhui; Yin, Haidi et al. (2014) Analysis of glycan variation on glycoproteins from serum by the reverse lectin-based ELISA assay. J Proteome Res 13:2197-204
Thakolwiboon, Smathorn; Zhu, Jianhui; Liang, Qixing et al. (2014) Heterogeneity of The CD90(+) Population in Different Stages of Hepatocarcinogenesis. J Proteomics Bioinform 7:296-302
Zhu, Jianhui; Lin, Zhenxin; Wu, Jing et al. (2014) Analysis of serum haptoglobin fucosylation in hepatocellular carcinoma and liver cirrhosis of different etiologies. J Proteome Res 13:2986-97

Showing the most recent 10 out of 12 publications