Cancer Letters

Cancer Letters

Volume 469, 28 January 2020, Pages 68-77
Cancer Letters

Original Articles
CircRNA cRAPGEF5 inhibits the growth and metastasis of renal cell carcinoma via the miR-27a-3p/TXNIP pathway

https://doi.org/10.1016/j.canlet.2019.10.017Get rights and content

Highlights

  • cRAPGEF5 downregulation is associated with RCC progression and predicts poor prognosis in RCC.

  • cRAPGEF5 inhibits RCC growth and metastasis by sponging miR-27a-3p.

  • miR-27a-3p facilitates RCC proliferation and migration by regulating TXNIP.

  • cRAPGEF5 suppresses RCC by the miR-27a-3p/TXNIP pathway.

Abstract

Circular RNAs (circRNAs) are reported to act as important regulators in cancers. CircRNA RAPGEF5 (cRAPGEF5) is derived from exons 2–6 of the RAPGEF5 gene and may promote papillary thyroid cancer progression. However, the role of cRAPGEF5 in renal cell carcinoma (RCC) remains unclear. In this study, we found cRAPGEF5 to be significantly downregulated in RCC tissues. Among 245 RCC cases, cRAPGEF5 downregulation correlated positively with aggressive clinical characteristics and independently predicted poor overall survival and recurrence-free survival. Functional assays demonstrated that cRAPGEF5 suppresses RCC proliferation and migration in vitro and in vivo. Mechanistically, RNA Immunoprecipitation and circRNA in vivo precipitation assays showed that cRAPGEF5 functions as a sponge of oncogenic miR-27a-3p, which targets the suppressor gene TXNIP. Interactions between miR-27a-3p and cRAPGEF5 or TXNIP were confirmed by dual-luciferase reporter assays. In conclusion, cRAPGEF5 plays a role in suppressing RCC via the miR-27a-3p/TXNIP pathway and may serve as a promising prognostic biomarker and novel therapeutic target for RCC patients.

Introduction

The incidence of renal cell carcinoma (RCC) has been increasing, with more than 400,000 estimated new cases and more than 170,000 deaths in 2018 worldwide [1]. As the most common type of kidney cancer, clear cell RCC (ccRCC) accounts for approximately 70–75% of cases [2]. Previous studies have reported that approximately 20–30% of RCC patients present with distant metastases at primary diagnosis, and nearly 25% of primary nonmetastatic cases treated with nephrectomy developed metastasis [3,4]. Although considerable improvements in the 5-year relative survival rates have been reported, the overall prognosis is far from satisfactory, particularly for metastatic RCC [5]. Thus, the potential mechanisms related to the development and progression of RCC still need to be further explored, and novel reliable therapeutic targets and predictive biomarkers are also needed to improve the survival rate of RCC patients.

Circular RNA (circRNA) is a novel class of endogenous RNA and uniquely presents with covalently closed loop structures [6], which is different from linear mRNA or long noncoding RNA. circRNAs are derived from precursor mRNAs through backsplicing and lack 5′ caps and 3’ polyadenylated tails [6,7]. Many studies have reported that circRNAs are widely expressed in a variety of mammalian tissues or cells and display specific expression patterns according to different cell types, tissues and developmental stages [[8], [9], [10], [11]]. Recently, a series of studies demonstrated that circRNAs were not byproducts of error splicing but were functional regulators [8] and were associated with many cancer types [12], including lung cancer [13], prostate cancer [14], breast cancer [15], glioma [16], hepatocellular carcinoma (HCC) [17,18], and renal cell carcinoma [19].

miRNAs are small noncoding RNAs and function mainly by regulating the expression of target genes [20,21]. Considerable miRNAs have been reported to be involved in the carcinogenesis and progression of RCC [22,23]. Recent studies found that circRNAs could function as sponges of miRNAs and regulate gene expression by blocking the activities of miRNAs [24]. Previous sequencing data show that hsa_circ_0001681 (derived from the RAPGEF5 gene, termed cRAPGEF5) is abnormally expressed in various cancers, including glioblastoma [25], liver cancer [26] and thyroid cancer [27]. A recent study reported that cRAPGEF5 could promote the proliferation and metastasis of papillary thyroid cancer through the miR-198/FGFR1 pathway [28]. However, whether cRAPGEF5 is abnormally expressed and plays a regulatory role in RCC remains unclear.

In the present study, we examined cRAPGEF5 expression in RCC tissues and identified that cRAPGEF5 is downregulated in RCC tissues and related to RCC prognosis. We further explored the functions and mechanisms of cRAPGEF5 in RCC. Functionally, silencing cRAPGEF5 enhances the growth, migration, and invasion abilities of RCC cells. Mechanistically, cRAPGEF5 could inhibit the progression of RCC by acting as a sponge of oncogenic miR-27a-3p to upregulate TXNIP. Therefore, cRAPGEF5 may serve as a novel therapeutic target and predictive biomarker for RCC patients.

Section snippets

Patients and samples

A total of 42 pairs of RCC and adjacent nontumor tissues and a cohort of 245 RCC tumor tissues were obtained from surgical resection specimens of RCC patients at Changzheng Hospital (Shanghai, China). The samples were quickly frozen in liquid nitrogen after surgery and then stored at the Biobank of Shanghai Changzheng Hospital until use. All samples were diagnosed by two pathologists independently. None of the patients underwent preoperative treatment. Written informed consent was obtained from

Expression and circRNA characterization of cRAPGEF5 in RCC

Bioinformatics analysis showed that cRAPGEF5 is an exonic circRNA that is cyclized with the second through sixth exons of the RAPGEF5 gene (Fig. 1A) [29]. A previous study found that cRAPGEF5 is associated with the progression of papillary thyroid carcinoma [28]; however, the role of cRAPGEF5 in RCC remains unclear. To measure the expression level of cRAPGEF5 in RCC tissues, we first designed specific PCR primers for cRAPGEF5, which were verified through Sanger sequencing and agarose gel

Discussion

The function of circRNA in tumors has received increasing attention, but its role in RCC tumorigenesis and progression remains unclear. In our study, the level of cRAPGEF5 in RCC was found to be relatively low; however, this result is different from the results in thyroid cancer [28], probably due to the tissue-specific expression of cRAPGEF5. Moreover, due to the limitation of the number of cases in this study, more cases may be required to fully support this conclusion.

The regulatory

Declaration of competing interest

All of the contributors in this work declared no conflicts of interest.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (81730073, 81874093, 81874093 and 81902560) and the Shanghai Sailing Program (19YF1448300).

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    These authors contributed equally to this work.

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