Abstract B20: Simultaneous measurement of 92 serum protein biomarkers for the development of a multiprotein classifier for ovarian cancer detection
Authors: Skubitz Amy P.N. , Boylan Kristin L.M. , Geschwind Kate , Cao Qing , Starr Timothy K. , Geller Melissa A. , Bast Robert C. , Lu Karen H. , Koopmeiners Joseph S. 
Published: August 2018

Abstract
Currently, there are no FDA-approved screening tools for detecting ovarian cancer in the general population. The two most common ovarian cancer biomarkers, CA125 and HE4, when used individually are neither adequately sensitive nor specific enough to be used for screening. We hypothesize that by using a combination of biomarkers for screening, it will be possible to increase the sensitivity and specificity over CA125 alone. Development of a blood-based assay for ovarian cancer detection could significantly improve the survival of patients if it identified cancers earlier. In this study, we used Proseek Multiplex Oncology II plates to simultaneously measure the expression of 92 cancer-related proteins in serum using proximity extension assays. This technology combines the sensitivity of the polymerase chain reaction with the specificity of antibody-based detection methods, allowing multiplex biomarker detection and high-throughput quantification. In a review of the literature, we found that 44 of these 92 proteins had been studied individually and were present at elevated levels in ovarian cancer samples (e.g., serum or tissues). This study is the first to examine the levels of 92 proteins simultaneously using a large cohort of ovarian cancer serum samples. We analyzed one microliter of serum from each of 60 women with late stage (Stages III and IV) high-grade serous ovarian cancer and compared the values obtained to those from 241 age-matched healthy women. The data from these 301 serum samples were normalized across four Proseek Multiplex Oncology II 96-well plates and across the different sources of sera. Principal component analysis and unsupervised hierarchical clustering separated patients into the two major groups of ovarian cancer and healthy, with minimal misclassification. Data from the Proseek plates for CA125 levels exhibited a strong correlation with previously measured clinical values for CA125 (correlation coefficient of 0.90, with a 95% confidence interval of 0.88-0.92). CA125 and HE4 were detected at low levels in healthy samples, while higher levels were found in the ovarian cancer cases. Overall, we identified 45 proteins that showed a significant difference (p &lt; 0.001) between ovarian cancer and healthy samples, many of which could serve as novel serum biomarkers for ovarian cancer. In total, 46 proteins had an estimated area under the ROC curve (AUC) of 0.7 or greater. CA125 alone achieved a sensitivity of 0.92 and a specificity of 0.98. In addition, we generated a multiprotein classifier using 10 proteins (including CA125) that increased the sensitivity to 0.97, while holding the specificity fixed at 0.98. This 5% increase in sensitivity will have a significant effect on the number of women correctly identified when screening a large population. Our data demonstrate that the Proseek technology can replicate the results established by conventional clinical assays for known biomarkers, identify new candidate biomarkers, and improve the sensitivity and specificity of CA125 alone. Additional studies using a larger cohort of early-stage ovarian cancer patients will allow for validation of these biomarkers and lead to the development of a screening tool for detecting early-stage ovarian cancer in the general population.
Citation Format: Amy P.N. Skubitz, Kristin L.M. Boylan, Kate Geschwind, Qing Cao, Timothy K. Starr, Melissa A. Geller, Robert C. Bast, Jr., Karen H. Lu, Joseph S. Koopmeiners. Simultaneous measurement of 92 serum protein biomarkers for the development of a multiprotein classifier for ovarian cancer detection. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr B20.