HE4 and epithelial ovarian cancer: Comparison and clinical evaluation of two immunoassays and a combination algorithm☆
Introduction
Ovarian cancer is the sixth most common cancer in women in the European Union, with an incidence of 18/100,000 women and represents the first cause of death for gynecological cancer and the third cause of death for oncologic diseases [1]. In Italy the standardized incidence, according to the National Cancer Register, is 13.5/100,000 and about 3000 deaths are reported every year, with a standardized mortality of 4.5/100,000 [2]. As for other malignancies, the survival rate is inversely proportional to the stage of the disease at diagnosis; women diagnosed in stages I–II have a 70% survival at 5 years, compared to less than 30% for patients in stages III and IV [1], [3], [4]. Thus, an early diagnosis in women presenting with suspect signs or symptoms is required in order to guarantee a better prognosis.
Laboratory diagnosis for ovarian cancer is mostly based on the quantitative determination of CA 125; though a broad array of immunoassays is available for the measurement of this mucin, established immunoassays for CA125 rely on the employ of well validated monoclonal antibodies. CA 125 is commonly employed for the baseline evaluation before treatment, during chemotherapy to evaluate the response and in patients’ monitoring during follow-up to detect recurrences earlier than the development of clinical symptoms [1], [4], [5], [6]. However the use of this marker as a first-line screening assay, alone or in combination with other procedures, is not recommended owing to the relatively low specificity and positive predictive value [7], [8]. Over the last years a number of additional markers for EOC have been proposed and studied, either alone or, most commonly, in association on the purpose of increasing the diagnostic accuracy, especially in the earlier stages of disease. In the majority of studies the human epididymis protein 4 (HE4) has emerged as one of the most promising markers [9], [10]. HE4 was initially identified in the epithelium of the distal epididymis and originally predicted to be a protease inhibitor involved in sperm maturation. It is also called WFDC2 because it contains two whey acid protein domains and a “four disulphide bond core” made up from eight cysteine residues.
The relevance of this marker in gynecological oncology is due to the fact that HE4 protein expression is highly restricted in normal tissue to the reproductive tracts and respiratory epithelium and is commonly overexpressed in epithelial ovarian cancer (EOC). While HE4 levels in healthy individuals have been reported to increase with age and are significantly higher in menopause [10], [11], [12], high levels are found in the serum of patients with EOC, mainly in serous and endometroid cancers [10], as well as in patients with endometrial cancer, but not in women with benign gynecological diseases or with endometriosis. As a consequence, HE4 has been reported to have a clear specificity edge over CA125 and also a better sensitivity for EOC, both in general and in the early stages as well as in the monitoring of treated patients [12], [13], [14], [15], [16].
The idea of combining the results of CA125 and HE4 in an algorithm (ROMA: Risk for Ovarian Malignancy Algorithm) that may help assessing the increased risk for EOC in symptomatic women has also been proposed [17], [18]. Furthermore, while the first studies on HE4 have been carried out with a conventional enzyme immunoassay (EIA), a fully automated chemiluminescent immunoassay has been recently made available. We therefore aimed at evaluating this new assay for HE4 in comparison with the EIA and also to verify the diagnostic accuracy for EOC of HE4 determination. We also evaluated the results of the ROMA algorithm generated by either HE4 immunoassay on a preselected population of Italian women.
Section snippets
Patients
In this retrospective study the population was selected in order to obtain a fairly balanced number of cases with benign or malignant ovarian lesions, an adequate representation of healthy controls and a proportion of pre- and post-menopausal women similar to that observed in routine clinical practice. Overall, 259 female patients were selected among those admitted to the Gynecologic Oncology unit of the University of Brescia from 2006. The inclusion criteria were: availability of complete
HE4 assays comparison
The intra-assay and interassay imprecision (CV%) of the CMIA HE4 assay was 2.11% and 3.64% on the low level control, 2.63% and 3.13% on the medium level control, 2.93% and 3.70% on the high level control. By comparison, the intraassay and interassays CVs% for the EIA HE4 assay, also evaluated on controls, ranged from 5.8% to 10.6% and from 6.8% to 10.3%, respectively.
For the CMIA assay, the average recovery on the two serially diluted specimens was 98.5 ± 9.0% (median: 98.5%) and the correlation
Discussion
Despite the severity of ovarian cancer, the low prevalence (about 40 cases per 100,000 in women aged more than 50 years) and the invasive nature of diagnostic procedures by laparoscopy or laparotomy is a serious hurdle for establishing screening policies. A 99.7% specificity will have anyway a positive predictive value of 10% at best, resulting in 9 healthy women undergoing invasive testing for each ovarian cancer diagnosed [4], [6], [7], [8] and in a recent experience in which CA125 has been
Acknowledgements
We wish to thank all the nurses working in the Department of Obstetrics and Gynaecology of the University of Brescia, and especially Ms. Margherita Franzoni, for the assistance in collecting blood samples. We are also grateful to Ms. Michela Faustini for her essential technical contribution in performing the ARCHITECT assays. We are also grateful to Dr. Michele Tronchin (Abbott Diagnostics, Italy) for having performed some of the statistical evaluations.
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Disclaimer: Dr. Claudio Galli is currently employed by Abbott Diagnostics as the Scientific Affairs Manager, Italy.