Abstract 542: CEACAM6 is upregulated by the H Pylori CAG-A oncoprotein, and is a candidate biomarker for the diagnosis of early gastric cancer

Abstract

Abstract Early detection of gastric cancers confers a significant mortality benefit, but remains a diagnostic challenge. We aim to identify and develop novel cell-surface biomarkers to improve the efficiency of gastric cancer detection by fluorescent endoscopy. Traditional approaches for surface biomarker discovery compare ‘normal’ and ‘malignant’ cell lines, but are prone to artifacts of cell line derivation and evolution. Our strategy exploits the dependence of gastric cancers on CAG-A, the Helicobacter Pylori oncoprotein. CAG-A incorporation is necessary and sufficient for gastric cancer pathogenesis, and is accordingly observed in early cancer specimens, but is intracellular and not expressed at high enough levels to be an applicable biomarker for cancer diagnosis. We therefore designed a strategy to screen for cell surface alterations induced by CAG-A overexpression, which may be retained in gastric cancers through oncogenic addiction. We used an inducible system for CAG-A expression in gastric MKN28 cells to compare the gene expression profile of cells with and without CAG-A overexpression, on the Affymetrix Gene 1.0 platform. We focused on putative membrane protein transcripts, to identify differentially upregulated candidates in CAG-A overexpressing cells. The top 10 hits from our screen were then analyzed in gene expression and RNAseq datasets from the Singapore Gastric Cancer Consortium, comparing the transcriptome of gastric cancer with adjacent normal tissue. Amongst our top hits in the in-vitro CAG-A screen, we noted a significant enrichment of the cell adhesion molecule CEACAM6 in gastric cancer samples when compared to normal tissue (∼8 fold increase in gene expression by microarray analysis (n = 100 normals, 200 cancers), and ∼200 fold increase in matched RNAseq analysis (n = 10 each)). We confirmed CEACAM6 to be upregulatd by CAG-A using RT-PCR on the inducible CAG-A system, and also in other gastric cell lines infected with H Pylori. Finally, we compared CEACAM6 protein expression on a gastric tissue microarray, using quantitative immunohistochemistry through automated spectral microscopy on the Vectra 2 platform. We confirm a 10-fold increase in CEACAM6 protein in gastric cancers as compared to normal tissue (n = 63 cancer, 52 normal). Fluorescently conjugated antibodies to these cell surface markers bind avidly to freshly resected xenografts (n = 3) of gastric cancer, without fixation. Together, these results suggest that CEACAM6 upregulation is a cell surface response to the introduction of H Pylori CAG-A in gastric epithelial cells, and is retained in a significant fraction of gastric cancers. It appears to be a promising candidate for a fluorescently conjugated biomarker to aid with the endoscopic diagnosis of early gastric cancer. Citation Format: Anand D. Jeyasekharan, Rony K. Roy, Michal M. Hoppe, Kar Tong Tan, Henry Yang, Patrick Tan, Phillip H. Koeffler, Khay Guan Yeoh, Singapore Gastric Cancer Consortium. CEACAM6 is upregulated by the H Pylori CAG-A oncoprotein, and is a candidate biomarker for the diagnosis of early gastric cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 542. doi:10.1158/1538-7445.AM2015-542