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BACKGROUND: The incidence of esophageal adenocarcinoma (EAC) has been rapidly increasing globally. The majority of EAC cases are diagnosed at late stages leading to poor 5-year survival of APPROACH & METHODOLOGY: Serum samples were collected from consenting patients undergoing endoscopy (Control - 16, BE - 13, and EAC - 10). The control group consisted of all patients not classified as BE with or without dysplasia, or EAC. LeMBA-MRM-MS was performed using 4 different lectins with different glycan specificities; AAL (Fucoseα1-2, -3, -6 linked glycan), EPHA (bisecting GlcNAc), JAC (Galα1-6GalNAc and Galβ1-3GalNAc), NPL (Manα1-6Man). Statistical analysis was performed using Shiny MixOmics as previously described [3]. RESULTS & DISCUSSION: Several lectin-protein candidate biomarkers cross-validated in the USA cohort. Most notably, differentially glycosylated complement component C9 is highly diagnostic in both cohorts, achieving area under the receiver operating curve (AUROC) of 0.74 (AAL-C9) to 0.90 (JAC-C9) as a single marker. Complement C9 binding to all four monitored lectins were increased 1.4 to 1.8 fold in EAC, compared to either BE or controls. Another consistent biomarker is gelsolin, with its binding to EPHA and NPL lectin significantly decreased in EAC, compared to either BE or controls. Hence, glycosylated C9 and gelsolin show promise as potential serum biomarkers for EAC screening, and will be further evaluated in prospective cohorts. REFERENCES: [1] Loo et al., J Proteome Res 9, 5496-5500 (2010); [2] Choi et al., Electrophoresis 32, 3564-3575 (2011); [3] Shah et al., Molecular & Cellular Proteomics 14:3023-39 (2015). Citation Format: Alok K. Shah, Virendra Joshi, Kim-Anh Le Cao, David C. Whiteman, Andrew P. Barbour, Michelle M. Hill. Towards a screening blood test for esophageal adenocarcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4942.

BACKGROUND: Esophageal adenocarcinoma (EAC) is one of the most rapidly increasing cancers globally. The majority of EAC cases are diagnosed at very late stages during pathogenesis hence APPROACH & METHODOLOGY: We focused on alterations in circulatory protein glycosylation, using a panel of 20 lectins to isolate different glycan structures on serum glycoproteins, as reported recently [1, 2]. Serum samples from healthy (n = 9), BE (n = 10) and EAC (n = 10) patient groups were analyzed by lectin magnetic bead array-coupled mass spectrometry (LeMBA-MS/MS) [1, 2]. Data analysis was performed using a customized database and analysis package “GlycoSelect” which incorporates outlier detection and sparse Partial Least Squares regression discriminant analysis (sPLS-DA) [3]. RESULTS & DISCUSSION: We identified a ranked list of candidate glycobiomarkers that distinguish a) EAC from BE b) BE from healthy and c) EAC from healthy group. In general, glycoproteins bound several lectins, reflecting heterogeneity and multiplicity of glycosylation. Specific glycan structure changes were observed as loss and gain of binding to a single lectin while maintaining binding to other lectins. Top two candidate biomarkers were validated using orthogonal validation technique LeMBA-immunoblotting in an independent patient cohort (n = 80). The biomarkers showed Area under Receiver Operating Characteristic curve (AUROC) of 0.74 to discriminate EAC from BE and 0.71 to discriminate BE from healthy patient group. Future work will validate all candidate protein-lectin pairs using lectin-affinity array coupled with triple quadrupole quantitative mass spectrometry measurements for the independent patient cohort. The specificity and sensitivity of panels of glycoprotein biomarkers will be determined for formulating a serum screening test for BE and EAC. [1] Choi et al., Electrophoresis 32, 3564-3575 (2011) [2] Loo et al., J Proteome Res 9, 5496-5500 (2010) [3] Lê Cao et al., BMC Bioinformatics 12, 253-268 (2011) Citation Format: Alok K. Shah, David Chen, Kim-Anh Le Cao, Eunju Choi, Derek Nancarrow, David Whiteman, Nicholas A. Saunders, Andrew P. Barbour, Michelle M. Hill. Discovery and validation of novel serum glycoprotein biomarkers for Barrett's esophagus and esophageal adenocarcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2492. doi:10.1158/1538-7445.AM2014-2492