Detection of Differential Levels of Proteins in the Urine of Patients with Endometrial Cancer: Analysis Using Two-Dimensional Gel Electrophoresis and O-Glycan Binding Lectin

Abstract

Cancers can cause some proteins to be aberrantly excreted or released in the urine, which can be used as biomarkers. To screen for potential biomarkers for endometrial cancer (ECa), the urinary proteins from patients who were newly diagnosed with early stage ECa and untreated controls were separated using two-dimensional gel electrophoresis (2-DE) and followed by image analysis. The altered levels of zinc alpha-2 glycoprotein, alpha 1-acid glycoprotein, and CD59 were detected in the patients compared to the controls. In addition, the urine of the ECa patients was also found to contain relatively lower levels of a fragment of nebulin when the 2-DE separated urinary proteins were probed using champedak galactose binding (CGB) lectin. The different levels of the nebulin fragment were further validated by subjecting the urinary protein samples to CGB lectin affinity chromatography and analysis of the bound fractions by LC-MS/MS. Our data is suggestive of the potential use of the differentially expressed urinary proteins as biomarkers for ECa although this requires further extensive validation on clinically representative populations.

Identification of O-glycosylated Proteins That Are Aberrantly Excreted in the Urine of Patients with Early Stage Ovarian Cancer

“…Data for the identification of the kininogen (KNG) spot cluster had been previously reported [7]. Analysis of the volumes of the proteins showed that the levels of LRG and CLU in the OCa patients were both 17-fold higher than the controls, while the level of the patients’ KNG was six-fold lower than the controls (Table 2).…”

“…CGB lectin was purified as previously described [7,8]. Briefly, crude extracts of the seeds of champedak were prepared by stirring of homogenized seeds in PBS, pH 7.4, overnight, and subjecting the supernatant to 60% ammonium sulfate precipitation.…”

“…In-gel digestion of protein plugs from 2-DE gels was carried out according to a previously described method [22]. On the other hand, on-membrane digestion was performed according to the method described by Mu et al [7]. The MALDI mass spectrometry was performed using the Applied Biosystem 4800 Proteomics Analyzer.…”

“…The attached O -glycans greatly influence the stability and tertiary structures of the glycoproteins and, hence, their functions in the biological system. Many serum O -glycosylated proteins have been previously implicated in cancer [5–7], either in the form of their altered O -glycan structures or their levels. These findings not only provide meaningful information about cancer development, but the glycoproteins that are modified may also be exploited for their potential use as cancer biomarkers.…”

Early biomarker discovery in urine of Walker 256 subcutaneous rat model

“…Bladder [21],Ovarian [22] Urine, Blood P02764 P02763 Alpha-1-acid glycoprotein (A1AG) Yes endometrial [23], bladder [24], lung [25] Urine, blood not peer-reviewed) is the author/funder. All rights reserved.…”