Clinical value of lncRNA CCAT1 in serum extracellular vesicles as a potential biomarker for gastric cancer

Abstract

Long non-coding RNAs (lncRNAs) in extracellular vesicles (EVs) are considered to be novel non-invasive biomarkers for gastric cancer (GC). lncRNA colon cancer-associated transcript 1 (CCAT1) is aberrantly expressed in certain types of cancer. However, the role of EV lncRNA CCAT1 in patients with GC remains unclear. The current study aimed to assess the expression levels of lncRNA CCAT1 in the serum EVs of patients with GC and evaluate its potential clinical value. EVs were isolated from serum using a commercial kit and ultracentrifugation, and were identified by transmission electron microscopy, nanoparticle tracking analysis and western blotting. Serum EV lncRNA CCAT1 levels in patients with GC, chronic gastritis or atypical hyperplasia and healthy control subjects were detected by reverse transcription-quantitative PCR. Additionally, lncRNA CCAT1 was detected in GC and adjacent non-cancerous tissue samples. Serum EVs were successfully isolated and identified in all patients. The results revealed that serum EV lncRNA CCAT1 levels in patients with GC were significantly higher compared with those in healthy controls, patients with chronic gastritis or atypical hyperplasia (all P<0.05). Additionally, EV lncRNA CCAT1 expression levels were significantly different among various groups based on the depth of invasion, distant metastasis and the Tumor-Node-Metastasis stage. The area under the curve (AUC) value of EV lncRNA CCAT1 was 0.890 [95% confidence interval (CI), 0.826-0.937] with 79.6% sensitivity and 92.6% specificity. The combination of EV lncRNA CCAT1 and carcinoembryonic antibody produced an AUC value of 0.910 (95% CI, 0.849-0.951) with the sensitivity and specificity of 80.5 and 92.6%, respectively. In addition, lncRNA CCAT1 was determined to be stable in serum EVs. The expression levels of lncRNA CCAT1 in GC tissue were positively correlated with those in serum EVs, and high levels of lncRNA CCAT1 were associated with a low disease-free survival rate in patients with GC. The results of the present study demonstrated that serum EV lncRNA CCAT1 levels were upregulated in patients with GC compared with those healthy subjects and patients with other illnesses, and may therefore be used as a novel biomarker for this type of cancer.

Keywords: biomarker; colon cancer-associated transcript 1; extracellular vesicle; gastric cancer; long non-coding RNA.

Figure 1.

Characterization of serum EVs. EVs were isolated and characterized from human serum using a commercial kit and ultracentrifugation. (A) Transmission electron microscopy micrographs. Magnification, ×40,000. (B) The size distribution of EVs was determined using a particle tracking analyzer. (C) Western blot analysis of EV markers CD9, CD63, TSG101 and calnexin. EVs, extracellular vesicles; TSG101, tumor susceptibility gene 101.

Figure 2.

Serum levels of EV long non-coding RNA CCAT1 in patients with gastric cancer, atypical hyperplasia and chronic gastritis, as well as healthy control subjects. EV, extracellular vesicle; CCAT1, colon cancer-associated transcript 1.

Figure 3.

Diagnostic value of serum EV lncRNA CCAT1 in GC. (A) Expression of lncRNA CCAT1 in patients with GC and different Tumor-Node-Metastasis stages. lncRNA CCAT1 was normalized to GAPDH. (B) ROC curve analyzing the diagnostic value of EV lncRNA CCAT1 and CEA. EV, extracellular vesicle; lncRNA, long non-coding RNA; CCAT1, colon cancer-associated transcript 1.

Figure 4.

Prognostic value of CCAT1 in patients with gastric cancer. (A) Long non-coding RNA CCAT1 expression levels were significantly associated with disease-free survival. (B) No significant association was observed between CCAT1 expression levels and overall survival. CCAT1, colon cancer-associated transcript 1.