Synthesis and Secretion of the Ovarian Cancer Antigen CA 125 by the Human Cancer Cell Line NIH:OVCAR-3

Lloyd, Kenneth O.; Yin, Beatrice W.T.

doi:10.1159/000050600

Cited by 52 publications

(78 citation statements)

References 18 publications

(19 reference statements)

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“…It is unlikely, however, that many of these sites are used as the content of N-linked glycan chains in purified CA125 is very low (15). It is also interesting to note that the sequence contains numerous lysine and arginine residues that are remote from the postulated O-glycosylation sites, and this could explain the sensitivity of CA125 to trypsin digestion (16). Searching for conserved domains in the NCBI Blast site revealed the presence of six SEA domains in the deduced protein sequence.…”

Section: Resultsmentioning

confidence: 99%

“…Thus, no consensus emerged from these studies concerning the biochemical nature of this antigen. Recently, however, our studies have strongly indicated that CA125 is a typical mucin molecule with a high carbohydrate content and a preponderance of serineand threonine-linked (O-linked) glycan chains (15,16). Possibly because of the mucinous nature of CA125 its peptide moiety has been very difficult to clone.…”

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confidence: 95%

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Molecular Cloning of the CA125 Ovarian Cancer Antigen

Yin

Lloyd

2001

Journal of Biological Chemistry

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549

404

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CA125 is an ovarian cancer antigen that is the basis for a widely used serum assay for the monitoring of patients with ovarian cancer; however, detailed information on its biochemical and molecular nature is lacking. We now report the isolation of a long, but partial, cDNA that corresponds to the CA125 antigen. A rabbit polyclonal antibody produced to purified CA125 antigen was used to screen a ZAP cDNA library from OVCAR-3 cells in Escherichia coli. The longest insert from the 54 positive isolated clones had a 5797-base pair sequence containing a stop codon and a poly(A) sequence but no clear 5 initiation sequence. The deduced amino acid sequence has many of the attributes of a mucin molecule and was designated CA125/MUC16 (gene MUC16). These features include a high serine, threonine, and proline content in an N-terminal region of nine partially conserved tandem repeats (156 amino acids each) and a C-terminal region non-tandem repeat sequence containing a possible transmembrane region and a potential tyrosine phosphorylation site. Northern blotting showed that the level of MUC16 mRNA correlated with the expression of CA125 in a panel of cell lines. The molecular cloning of the CA125 antigen will lead to a better understanding of its role in ovarian cancer.

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Section: Resultsmentioning

confidence: 99%

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confidence: 95%

Molecular Cloning of the CA125 Ovarian Cancer Antigen

Yin

Lloyd

2001

Journal of Biological Chemistry

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549

404

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“…The serum CA 125 antigen concentration is the 'gold standard' for ovarian cancer tumour markers (Lloyd and Yin, 2001). Although this protein was identified 20 years ago its structure and function has not been elucidated .…”

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confidence: 99%

Proteomic-based identification of haptoglobin-1 precursor as a novel circulating biomarker of ovarian cancer

et al. 2004

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Screening for specific biomarkers of early-stage detection of ovarian cancer is a major health priority due to the asymptomatic nature and poor survival characteristic of the disease. We utilised two-dimensional gel electrophoresis (2DE) to identify differentially expressed proteins in the serum of ovarian cancer patients that may be useful as biomarkers of this disease. In this study, 38 ovarian cancer patients at different pathological grades (grade 1 (n ¼ 6), grade 2 (n ¼ 8) and grade 3 (n ¼ 24)) were compared to a control group of eight healthy women. Serum samples were treated with a mixture of Affigel-Blue and protein A (5 : 1) for 1 h to remove high abundance protein (e.g. immunoglobulin and albumin) and were displayed using 11 cm, pH 4 -7 isoelectric focusing strips for the first dimension and 10% acrylamide gel electrophoresis for the second dimension. Protein spots were visualised by SYPRO-Ruby staining, imaged by FX-imager and compared and analysed by PDQuest software. A total of 24 serum proteins were differentially expressed in grade 1 (Po0.05), 31 in grade 2 (Po0.05) and 25 in grade 3 (Po0.05) ovarian cancer patients. Six of the protein spots that were significantly upregulated in all groups of ovarian cancer patients were identified by nano-electrospray quadrupole quadrupole time-of-flight mass spectrometry (n-ESIQ(q)TOFMS) and matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOFMS) as isoforms of haptoglobin-1 precursor (HAP1), a liver glycoprotein present in human serum. Further identification of the spots at different pathological grades was confirmed by Western blotting using monoclonal antibody against a haptoglobin epitope contained within HAP1. Immunohistochemical localisation of HAP1-like activity was present in malignant ovarian epithelium and stroma but strong immunostaining was present in blood vessels, areas with myxomatous stroma and vascular spaces. No tissue localisation of HAP1-like immunoreactivity was observed in normal ovarian surface epithelium. These data highlight the need to assess circulating concentration of HAP1 in the serum of ovarian cancer patients and evaluate its potential as a biomarker in the early diagnosis of ovarian cancer.

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“…CA125 is both N-and O-glycosylated in its N-terminal extracellular domain (Zurawski et al, 1988;Nagata et al, 1991;Lloyd et al, 1997;Lloyd and Yin, 2001), which is composed of a stalk domain next to the transmembrane span, more than 60 repeat structures (each of which consists of 156 amino acids) and an N-terminal extension (O'Brien et al, 2001;Yin and Lloyd, 2001). Towards the C-terminus, CA125 contains a putative transmembrane span and a short cytoplasmic tail (O'Brien et al, 2001).…”

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confidence: 99%

The cancer antigen CA125 represents a novel counter receptor for galectin-1

Seelenmeyer¹,

Wegehingel²,

Lechner³

et al. 2003

Journal of Cell Science

131

123

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CA125 is an ovarian cancer antigen whose recently elucidated primary structure suggests that CA125 is a giant mucin-like glycoprotein present on the cell surface of tumor cells. Here, we establish a functional link between CA125 and β-galactoside-binding, cell-surface lectins, which are components of the extracellular matrix implicated in the regulation of cell adhesion, apoptosis, cell proliferation and tumor progression. On the basis of mass spectrometry and immunological analyses, we find that CA125 is a counter receptor for galectin-1, as both soluble and membrane-associated fragments of CA125 derived from HeLa cell lysates are shown to bind specifically to human galectin-1 with high efficiency. This interaction is demonstrated (1) to depend on β-galactose-terminated, O-linked oligosaccharide chains of CA125, (2) to be preferential for galectin-1 versus galectin-3 and (3) to be regulated by the cellular background in which CA125 is expressed. Despite lacking a conventional signal peptide, a CA125 C-terminal fragment of 1148 amino acids, representing less than 10% of the full-length protein, retains the ability to integrate into secretory membranes such as the endoplasmic reticulum (ER) and the Golgi, and is targeted to the plasma membrane by conventional secretory transport. As demonstrated by a novel assay that reconstitutes non-conventional secretion of galectin-1 based on fluorescence-activated cell sorting (FACS), we find that tumor-derived HeLa cells expressing endogenous CA125 present more than ten times as much galectin-1 on their surface compared with non-tumor-derived, CA125-deficient CHO cells. Intriguingly, both the galectin-1 expression level and the cell-surface binding capacity for galectin-1 are shown to be similar in CHO and HeLa cells, suggesting that CA125 might be a factor involved in the regulation of galectin-1 export to the cell surface.

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Synthesis and Secretion of the Ovarian Cancer Antigen CA 125 by the Human Cancer Cell Line NIH:OVCAR-3

Cited by 52 publications

References 18 publications

Molecular Cloning of the CA125 Ovarian Cancer Antigen

Molecular Cloning of the CA125 Ovarian Cancer Antigen

Proteomic-based identification of haptoglobin-1 precursor as a novel circulating biomarker of ovarian cancer

The cancer antigen CA125 represents a novel counter receptor for galectin-1

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