Elsevier

Clinica Chimica Acta

Volume 501, February 2020, Pages 252-257
Clinica Chimica Acta

Exosomal long noncoding RNA lnc-GNAQ-6:1 may serve as a diagnostic marker for gastric cancer

https://doi.org/10.1016/j.cca.2019.10.047Get rights and content

Highlights

  • The expression of lnc-GNAQ-6:1 was significantly down-regulated in serum of patients with gastric cancer.

  • Exosomal long noncoding RNA lnc-GNAQ-6:1 may be a new tumor marker for gastric cancer diagnosis.

  • The higher purity exosomes can be isolated from the sera via this commercial exosome extraction kit.

Abstract

Background

Gastric cancer (GC) is one of the most aggressive cancers, with limited early diagnostic measures. Tumor-originated exosomal molecules are regarded as suitable candidates for non-invasive diagnosis. This study aimed to investigate the capacity of exosomal long noncoding RNA lnc-GNAQ-6:1 as a biomarker for early diagnosis of GC.

Methods

In this study, we collected sera from 43 patients with gastric cancer and 27 healthy subjects, then exosomes were isolated using commercial kits. Particle size analysis, Western bloting and protein-based exosomes quantification were conducted to identify the isolated exosomes and to evaluate its yield and purity. Expression levels of exosomal lnc-GNAQ-6:1 were detected by quantitative reverse transcription PCR (qRT-PCR). The serum concentrations of traditional biomarker (CA72-4, CEA, and CA19-9) were measured via a chemiluminescent detection system.The receiver operating characteristic curve (ROC) and area under curve (AUC) were used to estimate the diagnostic capacity. Furthermore, we analyzed the potential relationship between serum exosomal lnc-GNAQ-6:1 expression and clinicopathological parameters of gastric cancer.

Results

The exosomes extracted in this study exhibited the typical exosome characteristics and purity. Patients with gastric cancer had the higher exosome yield than healthy volunteer. The results of qRT-PCR showed that compared with the healthy control, the expression of lnc-GNAQ-6:1 was significantly lower in the gastric cancer group. The area under the ROC curve is 0.732, which was higher than the diagnostic accuracy of CEA, CA 19–9 and CA72–4. However, the expression level of lnc-GNAQ-6:1 was not correlated with gender, age, tumor metastasis, serum carcinoembryonic antigen (CEA), carbohydrate antigen 19–9 (CA19-9), and carbohydrate antigen 72–4(CA72-4).

Conclusions

Our data demonstrated that serum exosomal lnc-GNAQ-6:1 is lowly expressed in patient with gastric cancer and might be evaluated in larger studies as a new diagnostic marker for gastric cancer.

Introduction

Gastric cancer (GC) is the fifth most common cancer [1],and the second leading cause of cancer-related death in the world [2], [3], with 720,000 deaths occurred from above 950,000 new GC cases annually [4], [5], [6]. GC is a multi-step carcinogenic process and most patients are diagnosed at advanced stage due to the limited early diagnosis, which results in the lower survival rate [7], [8].Therefore, it is urgent to identify novel biomarker for patients with gastric cancer, so as to achieve early diagnosis and treatment.

Long noncoding RNA (LncRNA) are a group of noncoding RNAs with more than 200 nucleotides in length [9] and without protein coding ability, but occupies a majority of mammalian genome production [10], [11]. Increasing evidences have demonstrated that many lncRNAs are participated in the regulation of gene transcription and translation, cell growth, cell differentiation and tumorigenesis [12], [13], [14]. For example, lncRNA XIST were found markedly increased in GC and promote malignant phenotype of GC cell lines in vitro and in vivo [15]. GAS5 was found to be down-regulated in GC and GAS5′s ectopic expression could decrease gastric cancer cell proliferation and induce apoptosis in vitro and in vivo [16]. In addition, Shao et al [17] and Wang et al [18] revealed that different expression of lncRNAs correlates with the carcinogenesis and progression of GC and that lncRNAs could serve as early biomarkers or as treatment targets for GC.

Accumulating evidence has confirmed that tumor cell-derived ncRNAs (mRNAs, microRNAs and lncRNAs) can be secreted into body fluids such as blood, urine, milk, and saliva via exosomes [19], [20]. Exosomes, 30–100 nm sized membranous vesicles, are endogenously synthesized by all kinds of cell, which matters a lot in intercellular communication by transferring functional biomolecules (proteins, DNA, miRNA, lncRNA, and mRNA) to recipient cells [21], [22], [23]. Recent studies have demonstrated that the clinicopathological features of cancer can be reflected by the expression of specific exosomal lncRNAs, which means exosomal lncRNA might serve as novel tumor marker [24], [25], [26]. In light of these studies, we supposed that GC cells are able to generate and release exosomes and the examination of serum exosomal lncRNA may provide a noninvasive and convenient method for early screening and diagnosis of GC.

Based on our previous results of lncRNA microarray in GC cell-derived exosomes, lnc-GNAQ-6:1 was low expression in GC cell and exosomes compared with control. In this study, we identified the expression levels of lnc-GNAQ-6:1 in serum exosomes of patients with gastric cancer and healthy subjects. We also evaluated the potential correlation between the expression level of lnc-GNAQ-6:1 and clinicopathological features of patients with GC. Our data illustrated the potential of lnc-GNAQ-6:1 as a tumor marker for gastric cancer screening.

Section snippets

Patients

From April 2018 to July 2018, 43 patients with newly diagnosed gastric cancer admitted to the Affiliated Hospital of Xuzhou Medical University, 32 males, aged 43–77 years (median age 62 years),11 females, aged 42–81 years old (median age 66 years). Inclusion criteria: (1) All subjects were diagnosed by gastroscopy or histopathology; (2) no radiotherapy before surgery. Exclusion criteria: (1) combined with other cancer patients; (2) combined with other heart, kidney, liver dysfunction.

Characterization of exosomes in serum

To verify whether exosomes were extracted successfully, we identificated exosome by particle size analysis and western blot. Transmission electron microscopy directly observed exosomes, showing exogenous membrane structure is obvious, showing a typical tea tray structure (Fig. 1A). Particle size analysis showed that the average particle diameter was 60.1 nm (Fig. 1B), which was consistent with the range of exosomes reported in the literature. CD54 and TSG101, exosome protein markers, were

Discussion

The advantage of serological testing is that it not only reflects the local tissue of the tumor, but also has guiding significance for the whole cycle. The main methods of detecting gastric cancer are X-ray, gastroscope, CT, CEA, CA72-4 and CA19-9. X-ray has important diagnostic value for the detection of early gastric cancer, but the radiation is large. Gastroscope is the most reliable method for census of gastric cancer while it is invasive. The reliability of CT detection of early gastric

Conclusions

The results of the present study confirm that serum exosomal lnc-GNAQ-6:1 is lowly expressed in patient with gastric cancer and might be evaluated in larger studies as a new diagnostic marker for gastric cancer.

Acknowledgements

This work was supported by the grant from the Department of Science & Technology, Xuzhou, Jiangsu, China (No.KC17169), the Six Talent Peaks Project of Jiangsu Province of China (No.WSW-065) and the National Natural Science Foundation of China (No. 81802094).

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    These authors contributed equally: S. Li, M. Zhang, H. Zhang.

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