Abstract POSTER-CTRL-1211: Haptoglobin identified within ovarian cyst fluid as an accurate intraoperative diagnostic biomarker for epithelial ovarian cancer
Authors: Choolani Mahesh , Annamalai Loganath , Liu Lin , Razvi Khalil , Zhao Changqing , Rice Gregory , Mahyuddin Aniza P , Biswas Arijit , Chan Jerry , Kothandaraman Narasimhan 
Published: August 2015

Abstract
Introduction: Epithelial ovarian cancer (EOC) is the most lethal gynaecological malignancy, with a poor prognosis and low survival rate; most cases are diagnosed at a late stage due to the fact that symptoms at early stages are usually non-specific in nature. There is currently no screening method proven to be effective in improving the outcome of EOC patients; existing biomarkers for EOC, such as CA-125, generally suffer from a lack of specificity in early stage disease, which is the ideal time for therapeutic intervention.
Aim: We hypothesized that a single protein biomarker within the ovarian cyst fluid (OCF) could be identified, isolated, characterized and validated for application in a point-of-care device (POC), which could subsequently be used in operating theaters for triage for frozen section (FS).
Methods: We screened the OCF proteome by mass-spectrometry (MALTI-TOF/MS), confirmed the identity of the protein by western blot and SELDI immunocapture analysis. Next, demonstrated using tissue microarray (TMA) that cellular expression of haptoglobin varied in normal, benign and malignant ovarian tissue. We developed a simple ELISA and a rapid colorimetric assay that allowed semi-quantification of OCF haptoglobin intraoperatively. Finally, we validated a point-of-care test kit to accurately identify EOC using OCF with a higher predictive value than can be achieved using RMIs, and an equivalent accuracy to intraoperative FS.
Results: The OCF haptoglobin concentration in benign tumors was 0.70±0.09 mg/ml compared to 6.22±0.53 mg/ml and 6.57±0.65 mg/ml in early- and late-stage EOCs, respectively (P&lt;0.0001). We sourced a rapid colorimetric assay and determined the accuracy of this assay on archived OCF samples (n=124). The accuracy of this rapid colorimetric assay was similar to that of our in-house sandwich ELISA with sensitivity of 97.3% (95% CI: 84.2-99.9%), specificity of 92.0% (95% CI: 83.6-96.4%), PPV of 83.7% (95% CI: 68.7-92.7%) and NPV of 98.8% (95% CI: 92.4-99.9%). In an early phase clinical trial of the POC device, we prospectively tested 50 OCF samples real-time in the operation theater away from the surgeon’s view and the POC device tester blinded to the type of surgery performed and diagnosis. Visual calls from POC device were recorded and compared to histology. The POC device had an accuracy of 82% with sensitivity of 70% (95% CI: 35.1-91.9%), specificity of 85% (95% CI: 69.5-93.8%), PPV of 53.8% (95% CI: 26.1-79.6%), and NPV of 91.9% (95% CI: 77-97.9%).
Conclusion: This is the first study whereby an intraoperative tumor marker has been utilised in the differentiation between benign and malignant ovarian lesions. Its accuracy suggests that it could be utilised as a replacement or an adjunct to frozen section, particularly in situations where histopathological expertise is scarce.
Citation Format: Mahesh Choolani, Loganath Annamalai, Lin Liu, Khalil Razvi, Changqing Zhao, Gregory Rice, Aniza P Mahyuddin, Arijit Biswas, Jerry Chan, Narasimhan Kothandaraman. Haptoglobin identified within ovarian cyst fluid as an accurate intraoperative diagnostic biomarker for epithelial ovarian cancer [abstract]. In: Proceedings of the 10th Biennial Ovarian Cancer Research Symposium; Sep 8-9, 2014; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(16 Suppl):Abstract nr POSTER-CTRL-1211.