Quantitative Shotgun Proteomics Unveils Candidate Novel Esophageal Adenocarcinoma (EAC)-specific Proteins

Abstract

Esophageal cancer is the eighth most common cancer worldwide and the majority of patients have systemic disease at presentation. Esophageal adenocarcinoma (OAC), the predominant subtype in western countries, is largely resistant to current chemotherapy regimens. Selective markers are needed to enhance clinical staging and to allow targeted therapies yet there are minimal proteomic data on this cancer type. After histological review, lysates from OAC and matched normal esophageal and gastric samples from seven patients were subjected to LC MS/MS after tandem mass tag labeling and OFFGEL fractionation. Patient matched samples of OAC, normal esophagus, normal stomach, lymph node metastases and uninvolved lymph nodes were used from an additional 115 patients for verification of expression by immunohistochemistry (IHC).Over six thousand proteins were identified and quantified across samples. Quantitative reproducibility was excellent between technical replicates and a moderate correlation was seen across samples with the same histology. The quantitative accuracy was verified across the dynamic range for seven proteins by immunohistochemistry (IHC) on the originating tissues. Multiple novel tumor-specific candidates are proposed and EPCAM was verified by IHC.This shotgun proteomic study of OAC used a comparative quantitative approach to reveal proteins highly expressed in specific tissue types. Novel tumor-specific proteins are proposed and EPCAM was demonstrated to be specifically overexpressed in primary tumors and lymph node metastases compared with surrounding normal tissues. This candidate and others proposed in this study could be developed as tumor-specific targets for novel clinical staging and therapeutic approaches.

Distinct Microbiota Dysbiosis in Patients with Non-Erosive Reflux Disease and Esophageal Adenocarcinoma

“…Although our results were comparable with studies using similarly sized endoscopic biopsies [ 60 ], they were significantly fewer when compared with proteomic analysis using EAC surgical resections (>300 proteins) [ 30 , 61 ]. Small diagnostic mucosal biopsies at 3–5 mg [ 62 ] may not capture sufficient numbers of cancer cells for comprehensive proteomic analysis compared to much larger surgically resected tissue (30–60 mg) [ 61 ]. As a result, classical cancer markers such as epithelial cellular adhesion molecule (EpCAM) and inflammation markers were not identified in our EAC samples.…”

The Sequence-specific Peptide-binding Activity of the Protein Sulfide Isomerase AGR2 Directs Its Stable Binding to the Oncogenic Receptor EpCAM

“…In screening for cell lines that contain AGR2 and EpCAM ( Fig. 10 A ), we focused on the use of MCF7 cells because they express both proteins and have a wt-p53 pathway allowing for future impacts of AGR2 on p53 activity ( 27 ). Proximity Ligation assays were performed in MCF7 cells (AGR2+/EpCAM+) where the cells were incubated with different antibodies to the two proteins.…”

“…Our data highlight that the EpCAM receptor has specific AGR2 binding regions. The recent identification that EpCAM and AGR2 proteins are coexpressed at a very high frequency in human esophageal adenocarcinoma cancer biopsies ( 27 ) provides a clinical rationale to further dissect the AGR2-EpCAM pathway control in relation to carcinogenesis and potential therapeutics in this cancer type.…”

“…These data have implications for how this ER-resident chaperone mediates maturation of proteins destined for the plasma membrane. During these studies, a clinical proteomics analysis using resected biopsies from patients with esophageal adenocarcinoma identified AGR2 and EpCAM as highly expressed in primary adenocarcinoma as well as cancer-associated lymph nodes ( 27 ). This allows scientific information acquired on AGR2 and EpCAM pathway regulation to be translated into novel therapeutics that target the AGR2:EpCAM axis for improved cancer management.…”

Secretion of pro-oncogenic AGR2 protein in cancer

“…AGR2 was also found in other types of cancer that are related to hormone-dependence such as breast cancer [ 30 , 31 , 32 , 33 , 34 ], prostate cancer [ 35 , 36 , 37 ] and ovarian cancer [ 38 , 39 ]. Similarly, AGR2 expression was also found in non-hormone dependent cancers such as oesophageal cancer [ 17 , 40 ], gastric cancer [ 41 ], lung cancer [ 42 , 43 , 44 , 45 ] and head and neck cancer [ 46 ].…”