Expression of KOC, S100P, mesothelin and MUC1 in pancreatico-biliary adenocarcinomas: development and utility of a potential diagnostic immunohistochemistry panel

Abstract

Background: Pancreatico-biliary adenocarcinomas (PBA) have a poor prognosis. Diagnosis is usually achieved by imaging and/or endoscopy with confirmatory cytology. Cytological interpretation can be difficult especially in the setting of chronic pancreatitis/cholangitis. Immunohistochemistry (IHC) biomarkers could act as an adjunct to cytology to improve the diagnosis. Thus, we performed a meta-analysis and selected KOC, S100P, mesothelin and MUC1 for further validation in PBA resection specimens.

Methods: Tissue microarrays containing tumour and normal cores in a ratio of 3:2, from 99 surgically resected PBA patients, were used for IHC. IHC was performed on an automated platform using antibodies against KOC, S100P, mesothelin and MUC1. Tissue cores were scored for staining intensity and proportion of tissue stained using a Histoscore method (range, 0-300). Sensitivity and specificity for individual biomarkers, as well as biomarker panels, were determined with different cut-offs for positivity and compared by summary receiver operating characteristic (ROC) curve.

Results: The expression of all four biomarkers was high in PBA versus normal ducts, with a mean Histoscore of 150 vs. 0.4 for KOC, 165 vs. 0.3 for S100P, 115 vs. 0.5 for mesothelin and 200 vs. 14 for MUC1 (p < .0001 for all comparisons). Five cut-offs were carefully chosen for sensitivity/specificity analysis. Four of these cut-offs, namely 5%, 10% or 20% positive cells and Histoscore 20 were identified using ROC curve analysis and the fifth cut-off was moderate-strong staining intensity. Using 20% positive cells as a cut-off achieved higher sensitivity/specificity values: KOC 84%/100%; S100P 83%/100%; mesothelin 88%/92%; and MUC1 89%/63%. Analysis of a panel of KOC, S100P and mesothelin achieved 100% sensitivity and 99% specificity if at least 2 biomarkers were positive for 10% cut-off; and 100% sensitivity and specificity for 20% cut-off.

Conclusion: A biomarker panel of KOC, S100P and mesothelin with at least 2 biomarkers positive was found to be an optimum panel with both 10% and 20% cut-offs in resection specimens from patients with PBA.

Keywords: Biomarkers; Diagnosis; Immunohistochemistry; Pancreatic cancer.

Figure 1

Representative images of staining of all four biomarkers in normal tissue (normal pancreatic tissue) and range of staining intensities (weak, moderate and strong) in tumour tissue from tissue microarray cores.

Figure 2

ROC curves based on percentage of cells positive for any staining (weak, moderate or strong), in tumour and normal cases, for four biomarkers (A) KOC, (B) S100P, C) mesothelin and D) MUC1.

Figure 3

Sensitivity and specificity analysis of biomarkers for the diagnosis of pancreatico-biliary adenocarcinoma compared to normal tissue, based on five cut-offs for positivity: 5% positive cells of any staining intensity; 10% positive cells of any staining intensity; 20% positive cells of any staining intensity; 2 OR 3 intensity i.e. moderate or strong staining of cells; and Histoscore 20. Analysis is presented for A) KOC, B) S100P, C) mesothelin and D) MUC1.

Figure 4

*Combined Summary ROC curves for 10% (A) and 20% (B) cut-offs if only one biomarker was required to be positive in a panel. Four panels of biomarkers were compared. Panel 1 - KOC, S100P, Mesothelin and MUC1; Panel 2 - KOC, S100P, Mesothelin; Panel 3 - KOC, S100P; Panel 4 - KOC, Mesothelin. *Summary ROC curves plot sensitivity against specificity and draw a summary line depicting combined sensitivity and specificity of a panel. Combined Summary ROC curves compare different panels to show the most “accurate” panel. The summary line at the top left corner shows the biomarker which is most accurate compared to others lying lower and further to the right. This enables the most accurate panel to be identified.