Abstract
Gastric cancer is the third most common cause of death from cancer in the world and it remains difficult to cure in Western countries, primarily because most patients present with advanced disease. Currently, CEA, CA50 and CA72-4 are commonly used as tumor markers for gastric cancer by immunoassays. However, the drawback and conundrum of immunoassay are the unceasing problem in standardization of quality of antibodies and time/effort for the intensive production. Therefore, there is an urgent need for the development of a standardized assay to detect gastric cancer at the early stage. Aptamers are DNA or RNA oligonucleotides with structural domain which recognize ligands such as proteins with superior affinity and specificity when compared to antibodies. In this study, SELEX (Systematic Evolution of Ligands by Exponential enrichment) technique was adopted to screen a random 30mer RNA library for aptamers targeting CEA, CA50 and CA72-4 respectively. Combined with high-throughput sequencing, we identified 6 aptamers which specifically target for these three biomarkers of gastrointestinal cancer. Intriguingly, the predicted secondary structures of RNA aptamers from each antigen showed significant structural similarity, suggesting the structural recognition between the aptamers and the antigens. Moreover, we determined the dissociation constants of all the aptamers to their corresponding antigens by fluorescence spectroscopy, which further demonstrated high affinities between the aptamers and the antigens. In addition, immunostaining of gastric adenocarcinoma cell line AGS using CEA Aptamer probe showed positive fluorescent signal which proves the potential of the aptamer as a detection tool for gastric cancer. Furthermore, substantially decreased cell viability and growth were observed when human colorectal cell line LS-174T was transfected with each individual aptamers. Taking together, these novel RNA aptamers targeting gastrointestinal cancer biomarker CEA, CA50 and CA72-4 will aid further development and standardization of clinical diagnostic method with better sensitivity and specificity, and potentially future therapeutics development of gastric cancer.
Highlights
According to the statistics from World Health Organization 2015, Gastric cancer is the third most common cause of cancer-related death in the world [1]
Common biomarkers for gastrointestinal cancer can be largely classified as carcinoembryonic antigen (CEA), and tumor associated antigens, such as cancer antigen 19–9 (CA19-9), cancer antigen 50 (CA50) and cancer antigen 72–4 (CA72-4) [2]
Cancer antigen 72–4 (CA72-4), with a molecular weight of 220–400 kD, is a mucin-like high molecular weight tumor associated antigen, and it is considered as the first choice of tumor marker for gastric carcinoma because of a superior sensitivity than CEA and CA19-9 [1]
Summary
According to the statistics from World Health Organization 2015, Gastric cancer is the third most common cause of cancer-related death in the world [1]. Common biomarkers for gastrointestinal cancer can be largely classified as carcinoembryonic antigen (CEA), and tumor associated antigens, such as cancer antigen 19–9 (CA19-9), cancer antigen 50 (CA50) and cancer antigen 72–4 (CA72-4) [2]. Cancer antigen 50 (CA50), with a molecular weight of ~210 kD, is defined by the monoclonal antibody C 50 developed against a colorectal cancer (CRC) cell line COLO-205. It has elevated levels in serum and can be observed in a variety of malignancies, especially gastrointestinal cancers [4]. Cancer antigen 72–4 (CA72-4), with a molecular weight of 220–400 kD, is a mucin-like high molecular weight tumor associated antigen, and it is considered as the first choice of tumor marker for gastric carcinoma because of a superior sensitivity than CEA and CA19-9 [1]
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