Glycoproteomic markers of hepatocellular carcinoma‐mass spectrometry based approaches

Abstract

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. Cirrhosis is the final common pathway for several etiologies of liver disease, including hepatitis B and C, alcohol, and increasingly non‐alcoholic fatty liver disease. Only 20–30% of patients with HCC are eligible for curative therapy due primarily to inadequate early‐detection strategies. Reliable, accurate biomarkers for HCC early detection provide the highest likelihood of curative therapy and survival; however, current early‐detection methods that use abdominal ultrasound and serum alpha fetoprotein are inadequate due to poor adherence and limited sensitivity and specificity. There is an urgent need for convenient and highly accurate validated biomarkers for HCC early detection. The theme of this review is the development of new methods to discover glycoprotein‐based markers for detection of HCC with mass spectrometry approaches. We outline the non‐mass spectrometry based methods that have been used to discover HCC markers including immunoassays, capillary electrophoresis, 2‐D gel electrophoresis, and lectin‐FLISA assays. We describe the development and results of mass spectrometry‐based assays for glycan screening based on either MALDI‐MS or ESI analysis. These analyses might be based on the glycan content of serum or on glycan screening for target molecules from serum. We describe some of the specific markers that have been developed as a result, including for proteins such as Haptoglobin, Hemopexin, Kininogen, and others. We discuss the potential role for other technologies, including PGC chromatography and ion mobility, to separate isoforms of glycan markers. Analyses of glycopeptides based on new technologies and innovative softwares are described and also their potential role in discovery of markers of HCC. These technologies include new fragmentation methods such as EThcD and stepped HCD, which can identify large numbers of glycopeptide structures from serum. The key role of lectin extraction in various assays for intact glycopeptides or their truncated versions is also described, where various core‐fucosylated and hyperfucosylated glycopeptides have been identified as potential markers of HCC. Finally, we describe the role of LC‐MRMs or lectin‐FLISA MRMs as a means to validate these glycoprotein markers from patient samples. These technological advancements in mass spectrometry have the potential to lead to novel biomarkers to improve the early detection of HCC.

What Is Next in This “Age” of Heme-Driven Pathology and Protection by Hemopexin? An Update and Links with Iron

“…Additional monitoring of plasma proteins via glycan screening, including HPX [97], may also provide evidence of changes in liver function, thus aiding in the early detection and progress of HCC. This technique involves protein extraction using lectin-affinity binding of plasma samples.…”

Functional glycomics: Application to medical science and hepatology

“…Glycan analysis with mass spectrometry has provided insight in glyco‐biomarker research. For example, site‐specific glycopeptide analyses of hepatic glycoproteins using mass spectrometry yield a variety of glyco‐biomarkers for HCC and other liver diseases . Novel lectin‐based mass spectrometry analysis has also been developed .…”

“…As a result, one potential problem of using total N ‐glycan analyses as a marker of HCC and other liver diseases is that serum glycoprotein status is altered in many pathological conditions, rendering its prognostic value for a specific disease questionable. In the case of fucosylated AFP detection, mass spectrometry analysis yields better accuracy than for AFP; however, glycan analysis through mass spectrometry is difficult to use in a clinical setting due to its complicated and specialized equipment and cost.…”

“…51 Novel lectin-based mass spectrometry analysis has also been developed. 51 Combination of glyco-biomarkers, as well as conventional HCC markers, such as AFP and proteininduced vitamin K-deficient antagonist-II, results in much higher sensitivity and/or specificity. In addition, total Nglycan changes in serum glycoproteins have been proposed as a novel biomarker for HCC.…”

N-glycopeptide Signatures of IgA2 in Serum from Patients with Hepatitis B Virus-related Liver Diseases

“…Enhanced fucosylation of acute-phase proteins such as haptoglobin (11,12) have been reported in HCC. A recent review by Zhu et al summarized glycoproteomics markers of HCC especially based on mass spectrometry (MS) approaches (13).…”