Mass Spectrometric Assay for Analysis of Haptoglobin Fucosylation in Pancreatic Cancer

Abstract

A mass spectrometric method was developed to elucidate the N-glycan structures of serum glycoproteins and utilize fucosylated glycans as potential markers for pancreatic cancer. This assay was applied to haptoglobin in human serum where N-glycans derived from the serum of 16 pancreatic cancer patients were compared with those from 15 individuals with benign conditions (5 normals, 5 chronic pancreatitis, and 5 type II diabetes). This assay used only 10uL of serum where haptoglobin was extracted using a monoclonal antibody and quantitative permethylation was performed on desialylated N-glycans followed by MALDI-QIT-TOF MS analysis. Eight desialylated N-glycan structures of haptoglobin were identified where a bifucosylated tri-antennary structure was reported for the first time in pancreatic cancer samples. Both core and antennary fucosylation were elevated in pancreatic cancer samples compared to samples from benign conditions. Fucosylation degree indices were calculated and show a significant difference between pancreatic cancer patients of all stages and the benign conditions analyzed. This study demonstrates that a serum assay based on haptoglobin fucosylation patterns using mass spectrometric analysis may serve as a novel method for the diagnosis of pancreatic cancer.

Glycomic and Proteomic Profiling of Pancreatic Cyst Fluids Identifies Hyperfucosylated Lactosamines on the N-linked Glycans of Overexpressed Glycoproteins

“…Recent studies have reported that serum CA19-9 levels can detect pancreatic cancer independently (35) or as a component of a multiple marker panel (36), although other reports have concluded that it is not specific for the diagnosis of cancer (12,37). In addition to CA19-9, increases in core fucosylation of serum proteins such as ribonuclease 1 (38) and both core and outer arm fucosylation of haptoglobin (16,17) have been reported as putative biomarkers. A considerable amount of research has been conducted to investigate glycan biomarkers as a potential molecular marker system for pancreatic cancer; however, very little information is available with regard to the variety and abundance of glycans (or glycoproteins) that are present in the tissues and fluids of the pancreas.…”

“…A subgroup of the cysts, including several IPMN, was observed here to contain hyperfucosylated N-linked glycans. Increased fucosylation has continued to be implicated in the progression of cancer (13)(14)(15)(16), including specifically pancreatic cancer (16,17). As such, hyperfucosylated N-glycans and glycoproteins have been targeted for quantitative label-free bottom-up glycoproteomic analysis.…”

N-Glycosylation Is Critical for the Stability and Intracellular Trafficking of Glucose Transporter GLUT4

“…The results of the present study provide evidence that the N-glycan on GLUT4 not only plays a general role in protein stability, but also has a function in GLUT4 sorting and insulin responsiveness. It should be noted that several studies have demonstrated changes of N-glycan profiles under diabetic conditions (43)(44)(45)(46), and the effects of glycan profile alterations, especially in GLUT4, on the insulin resistance phenotype in type 2 diabetes should therefore be carefully investigated in future studies. Clarification of the key factors involved in N-glycan-related intracellular trafficking will be imperative for understanding the precise roles of the N-glycan functions in GLUT4.…”

Site-specific Glycoforms of Haptoglobin in Liver Cirrhosis and Hepatocellular Carcinoma

“…The authors also detected traces of doubly fucosylated Le(y)-type structures in tetra-antennary glycans. Lin et al (42) reported increased core fucosylation in patients with pancreatic cancer; both core and outer arm fucoses were detected among the doubly fucosylated glycans. Dimeric Lewis-A type (Le(a) on Le(a)) structures of Hp were found increased in colon cancer (43).…”