A multiplex platform for the identification of ovarian cancer biomarkers

Abstract

BackgroundCurrently, there are no FDA approved screening tools for detecting early stage ovarian cancer in the general population. Development of a biomarker-based assay for early detection would significantly improve the survival of ovarian cancer patients. MethodsWe used a multiplex approach to identify protein biomarkers for detecting early stage ovarian cancer. This new technology (Proseek® Multiplex Oncology Plates) can simultaneously measure the expression of 92 proteins in serum based on a proximity extension assay. We analyzed serum samples from 81 women representing healthy, benign pathology, early, and advanced stage serous ovarian cancer patients. ResultsPrinciple component analysis and unsupervised hierarchical clustering separated patients into cancer versus non-cancer subgroups. Data from the Proseek® plate for CA125 levels exhibited a strong correlation with current clinical assays for CA125 (correlation coefficient of 0.89, 95% CI 0.83, 0.93). CA125 and HE4 were present at very low levels in healthy controls and benign cases, while higher levels were found in early stage cases, with highest levels found in the advanced stage cases. Overall, significant trends were observed for 38 of the 92 proteins (p < 0.001), many of which are novel candidate serum biomarkers for ovarian cancer. The area under the ROC curve (AUC) for CA125 was 0.98 and the AUC for HE4 was 0.85 when comparing early stage ovarian cancer versus healthy controls. In total, 23 proteins had an estimated AUC of 0.7 or greater. Using a naïve Bayes classifier that combined 12 proteins, we improved the sensitivity corresponding to 95% specificity from 93 to 95% when compared to CA125 alone. Although small, a 2% increase would have a significant effect on the number of women correctly identified when screening a large population.ConclusionsThese 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 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12014-017-9169-6) contains supplementary material, which is available to authorized users.

Simultaneous Measurement of 92 Serum Protein Biomarkers for the Development of a Multiprotein Classifier for Ovarian Cancer Detection

“…In agreement with our previous study on the Oncology Iv2 plate (13) and others (25,28), we found high levels of both MK and IL6 in the sera of patients with ovarian cancer. FR-alpha, KLK11, and NECT4 also showed a significant increase in their levels of expression in the sera of ovarian cancer patients as previously reported (24,26,27).…”

“…The Proseek platform also includes three "Interplate controls" for data normalization between plates and three "Negative controls" to establish background levels. One microliter of serum from each of the samples was mixed with the Oncology II reagents following the manufacturer's protocol, then processed in combination with the Fluidigm BioMark HD high-throughput PCR instrument at the University of Minnesota Genomics Center as previously described (13). Data generated from the plates were analyzed, including normalization and linearization, per manufacturer's protocol.…”

“…Recently, we tested a small sample set (20 cases each of healthy, benign ovarian, early stage serous ovarian cancer, and late-stage serous ovarian cancer) on the Proseek Multiplex Oncology Iv2 panel to determine its feasibility as a means to identify candidate biomarkers for early stage serous ovarian cancer (13). We demonstrated that the Proseek technology provides similar results to conventional clinical assays for CA125, and can identify new candidate biomarkers that improve the sensitivity and specificity of CA125 alone.…”

High throughput proteomics identifies 484 high-accuracy plasma protein biomarker signatures for ovarian cancer

“…Our top-ranked model had a sensitivity of 0.85 and specificity of 0.91 under the same conditions. Similar to the results of these previous studies 8,9 , the performance of our models in the test-proportion of the discovery data is very good, with some models showing perfect classification. We also evaluated the selected models in two replication cohorts and found the performance similar, but somewhat lower than in the discovery set.…”

“…We then performed model selection as before based solely on the discovery data (benign tumours versus ovarian cancer stages III-IV) and identified a model consisted of 8 proteins. We finally added three proteins (WFDC2, KRT19 and FR-alpha) based on their previous association with ovarian cancer stages I-II in our modelling, or in previous literature 9,12,13 . The 11-protein panel consisted of MUCIN-16, SPINT1, TACSTD2, CLEC6A, ICOSLG, MSMB, PROK1, CDH3, WFDC2, KRT19 and FR-alpha.…”