Golgi protein 73 as a biomarker of hepatocellular cancer: Development of a quantitative serum assay and expression studies in hepatic and extrahepatic malignancies

In the search for serum markers of hepatocellular cancer, several investigators have recently focused on Golgi protein 73 [GP73; also known as Golgi membrane protein 1 (Golm1) or Golph2; Gen ID 51280]. GP73 is a 400– amino acid, 73-kD transmembrane glycoprotein that normally resides within the cis-Golgi complex. Its messenger RNA was first identified in a search for up-regulated hepatic genes in a patient with syncytial giant-cell hepatitis.1 Subsequent studies revealed minimal GP73 expression in normal hepatocytes but marked expression in patients with acute and chronic hepatitis and liver cirrhosis2,3 (Fig. 1A). This up-regulation occurred in response to viral and nonviral etiologies and thus appeared to be a general feature of liver disease. In addition to hepatocytes, GP73 was consistently expressed by normal biliary epithelial cells as well as hepatic stellate cells in injured livers.3 Further studies demonstrated constitutive expression in cells of the epithelial lineage, especially in the prostate, gut, breast, and thyroid, and within the central nervous system.1 Given its intracellular localization as a resident Golgi protein, its widespread expression in extrahepatic tissues, and its up-regulation in nonmalignant liver diseases, it came as a surprise when two reports identified a serum form of GP73 (sGP73) as a marker of hepatocellular cancer. Block and colleagues4 first demonstrated GP73 immunoreactivity in serum and found increased sGP73 levels in patients with hepatitis B virus–related hepatocellular cancer. In a subsequent study by Marrero et al.,5 sGP73 levels were significantly increased in patients with hepatitis C virus (HCV)–related hepatocellular carcinoma (HCC) in comparison with cirrhotic controls. The sensitivity and specificity of sGP73 for HCC were superior to those of alpha-fetoprotein, especially in early HCC. Similar results were reported in a Chinese study on patients with predominantly hepatitis B virus–related liver cancer.6 In response to these encouraging reports, GP73 was added to a group of emerging candidate HCC serum markers.7 The mechanism by which sGP73 reaches the circulation was worked out by Bachert and colleagues8 in cell culture studies. Despite its steady-state localization within the cis-Golgi complex, GP73 cycles through the distal secretary apparatus and transiently reaches the apical cell membrane, from which it returns to the Golgi complex via an endosomal retrieval pathway.9 A slightly smaller form of GP73 is present in the supernatants of several cell lines. This secreted form is generated by N-terminal cleavage of the molecule by the proprotein convertase furin after amino acid 55, resulting in the release of the large C-terminal ectodomain into the extracellular space8 (Fig. 1B). Using N-terminal sequencing, Gu and colleagues10 confirmed that the serum form of sGP73 is identical to the furin cleavage product identified in cell culture supernatants. This finding provides a mechanistic explanation for the appearance of sGP73 in serum. One drawback of the initial studies was that sGP73 was detected by western blotting, a semiquantitative and laborious test. sGP73 sandwich enzyme-linked immunosorbent assays (ELISAs) were first developed by Gary Norman at Inova Diagnostics and by Gu and colleagues.10 The assays were tested in two large patient cohorts from Italy and China, respectively. The results were disappointing. In the first study, sGP73 was found to be elevated in patients with liver disease but did not distinguish between HCC, cirrhosis, and chronic hepatitis.10 In the second study, which was reported in an abstract form, sGP73 was surprisingly found to be decreased in HCC patients.11 One potential methodological issue in both studies was the use of antibodies directed against bacterially expressed GP73. Given the extensive glycosylation and hyperfucosylation of GP73 in eukaryotic cells,12 these antibodies might not recognize the fully modified sGP73. Furthermore, recently described GP73-specific serum autoantibodies might interfere with the ELISA analysis.11 Abbreviations: aa, amino acid; ELISA, enzyme-linked immunosorbent assay; GP73, Golgi protein 73; HCC, hepatocellular carcinoma; HCV, hepatitis C virus; sGP73, serum form of Golgi protein 73; TM, transmembrane domain. Address reprint requests to: Claus J. Fimmel, Division of Gastroenterology, Hepatology, and Nutrition, Loyola University, 2160 South 1st Avenue, Maywood, IL 60153. E-mail: cfimmel@lumc.edu; fax: 708-216-4113. Copyright © 2009 by the American Association for the Study of Liver Diseases. Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hep.22944 Potential conflict of interest: Nothing to report.