Publications

Phase II biomarker trial of a multimarker diagnostic for ovarian cancer

Journal of Cancer Research and Clinical Oncology

Tracey Edgell, G. Martin-Roussety, G. Barker, D. J. Autelitano, D. Allen, P. Grant and G. E. Rice

Abstract

Purpose The primary hypothesis to be tested in this study was that the diagnostic performance (as assessed by the area under the receiver operator characteristic curve, AUC) of a multianalyte panel to correctly identify women with ovarian cancer was significantly greater than that for CA-125 alone.

Methods A retrospective, case–control study (phase II biomarker trial) was conducted that involved 362 plasma samples obtained from women with ovarian cancer (n = 150) and healthy controls (n = 212). A multivariate classification model was developed that incorporated five biomarkers of ovarian cancer, CA-125; C-reactive protein (CRP); serum amyloid A (SAA); interleukin 6 (IL-6); and interleukin 8 (IL-8) from a modelling cohort (n = 179). The performance of the model was evaluated using an independent validation cohort (n = 183) and compared with of CA-125 alone.

Results The AUC for the biomarker panel was significantly greater than the AUC for CA-125 alone for a validation cohort (p < 0.01) and an early stage disease cohort (i.e. Stages I and II; p < 0.01). At a threshold of 0.3, the sensitivity and specificity of the multianalyte panel were 94.1 and 91.3%, respectively, for the validation cohort and 92.3 and 91.3%, respectively for an early stage disease cohort.

Conclusions The use of a panel of plasma biomarkers for the identification of women with ovarian cancer delivers a significant increase in diagnostic performance when compared to the performance of CA-125 alone.

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Increased plasma concentrations of anterior gradient 2 protein are positively associated with ovarian cancer

Clinical Science

Tracey A. Edgell, Dong L. Barraclough, Antonio Rajic, Janu Dhulia, Kate J. Lewis, Jane E. Armes, Roger Barraclough, Philip S. Rudland, Gregory E. Rice and Dominic J. Autelitano

Abstract

Ovarian cancer is often asymptomatic and is diagnosed at an advanced stage with poor survival rates, thus, there is an urgent need to develop biomarkers for earlier detection of ovarian cancer. Here, we demonstrate for the first time that the previously-reported metastasis-inducing protein, anterior gradient protein 2 (AGR2), can be detected in the blood of ovarian cancer patients. Using a newly developed ELISA test, we show significantly increased concentrations of AGR2 protein in plasma from cancer patients relative to normal controls. Plasma AGR2 concentrations were highest in stage II and stage III ovarian cancer patients and were similarly elevated in patients with both serous and non-serous tumours. The identification of elevated plasma concentrations of AGR2 may provide a useful biomarker to aid in the discrimination of normal and ovarian cancer patients particularly when used in combination with CA125.

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Protein Depletion Using IgY from Chickens Immunised with Human Protein Cocktails

Preparative Biochemistry & Biotechnology

Antonio Rajic; Christiane Stehmann; Dominic J. Autelitano; Ana K. Vrkic; Christopher G. Hosking; Gregory E. Rice; Leodevico L. Ilag

Abstract

Given that proteomic analysis of complex protein mixtures may be restricted by the presence of highly abundant proteins, sample preparation to remove abundant proteins is essential for the analysis of low abundance proteins. Chickens are effective producers of antibodies (IgY) against mammalian proteins, able to produce large quantities of antibodies that can be recovered by simple non-intrusive extraction of egg yolk. The extraction procedure described uses a modification of the water dilution method (WD) to deplete lipids and lipoproteins followed by sequential precipitation with 31% ammonium sulphate and 12% poly ethylene glycol (PEG) producing IgY antibodies with greater than 95% purity and no loss in immunoreactivity. In the present study, various cocktails of the 12 most abundant human plasma proteins were used as immunogens to produce IgY antibodies. The anti-cocktail IgY antibodies were effectively used to sequentially and selectively immunodeplete abundant proteins from plasma. Also, affinity depletion (e.g., Affi-Gel Blue) was combined with immunodepletion to sequentially deplete abundant proteins from both plasma and urine. The current approach described allows the end user to mix and match sets of IgY cocktails to deplete tailored sets of targeted proteins dependent on their end use application.