Cancer Letters

Cancer Letters

Volume 423, 1 June 2018, Pages 113-126
Cancer Letters

Original Articles
Maintenance of stemness by miR-589-5p in hepatocellular carcinoma cells promotes chemoresistance via STAT3 signaling

https://doi.org/10.1016/j.canlet.2017.11.031Get rights and content

Highlights

  • miR-589-5p is significantly elevated in HCC tissues and cells.

  • Overexpression of miR-589-5p relates with poor overall and relapse-free survival of HCC patients.

  • miR-589-5p promotes stemness and chemoresistance in vitro and in vivo.

  • miR-589-5p promotes stemness and chemoresistance via activating STAT3 and signaling.

  • Recurrent gains contribute to miR-589-5p overexpression in HCC tissues.

Abstract

The strength and duration of STAT3 signaling are tightly controlled by multiple negative feedback mechanisms under physical conditions. However, how these serial feedback loops are simultaneously disrupted in cancers, leading to constitutive activation of STAT3 signaling in hepatocellular carcinoma (HCC), remains obscure. Here we report that miR-589-5p is elevated in HCC tissues, which is caused by recurrent gains. Overexpression of miR-589-5p correlates with poor overall and relapse-free survival in HCC patients. Upregulating miR-589-5p enhances spheroid formation ability, fraction of CD133 positive and side population cells, expression of cancer stem cell factors and the mitochondrial potential, and represses the apoptosis induced by doxorubicin in vitro and tumorigenicity in vivo in HCC cells; conversely, silencing miR-589-5p yields an opposite effect. Our findings further demonstrate miR-589-5p promotes the cancer stem cell characteristics and chemoresistance via targeting multiple negative regulators of STAT3 signaling pathway, including SOCS2, SOCS5, PTPN1 and PTPN11, leading to constitutive activation of STAT3 signaling. Collectively, our results unravel a novel mechanism by which miR-589-5p promotes the maintenance of stemness and chemoresistance in HCC, providing a potential rational registry of anti-miR-589-5p combining with conventional chemotherapy against HCC.

Introduction

Hepatocellular carcinoma (HCC) is the fifth most prevalent cancer and third-leading cause of cancer-related deaths worldwide due to heterogeity in clinical outcomes and biological activities [1]. Surgical resection is the first-line treatment for HCC, but unfortunately surgery is not applicable for the majority of HCC patients due to the advanced stages. Despite great progresses in systemic management and treatment in the recent years, the 5-year survival rate remains no more than 15%, which is largely attributed to the development of chemotherapeutic resistance [2]. The principal factor responsible for the failure of chemotherapy is the existence of cancer stem cells (CSCs) that are a minority cell population within tumors characterized by the capabilities of self-renewal and differentiation into the heterogeneous lineages of cancer cells which contributes to a hierarchical organization of cancer cells [3], [4], which is implicated in the progression and metastasis of various cancers [5], [6], [7]. Furthermore, CSCs have been reported to play an important role in the induction of chemotherapeutic resistance [8], [9], suggesting that these two cellular processes are intimately linked. Therefore, better understanding of the mechanism responsible for the maintenance of CSCs may facilitate to develop novel therapies aimed at eradicating the CSC population to achieve long-term remission and improve the survival rate in HCC patients.

Numerous studies indicated that constitutive activation of The Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway is constitutively activated in a number of human cancers, which contributes to the induction and maintenance of CSCs via transcriptionally regulating several well-known cancer stem cell factors, such as Nanog homeobox (NANOG) [10], [11]. The JAK/STAT was first identified as a critical regulator of cytokine signaling in a study of interferon signaling [12]. From a molecular vision, it is primarily consisted of three main components: a cell surface receptor JAK and two STAT proteins. The activation of JAK/STAT3 pathway starts with the binding of various ligands, such as interleukin-6 (IL-6), interferon (IFN), and IL-10, with JAK proteins. The activated JAKs further engage with cytokine receptors and phosphorylate tyrosine residues on the receptor, which further recruits and phosphorylated STATs. The phosphorylated STATs are released from the receptors and form homo- or hetero-dimers and translocate to the nucleus in which they further transcriptionally regulates downstream targeted genes [13]. Multiple liens of studies have reported that JAK/STAT signaling pathway was indispensable in many aspects of tumorigenesis, including proliferation, apoptosis, chemoresistance, and metastasis [14], [15]. Furthermore, numerous mechanisms have been reported to be involved in the aberrant activation of JAK/STAT signaling, including increased production of cytokines, and loss or decreased expression of negative regulators, such as suppressor of cytokine signaling (SOCS) or protein tyrosine phosphatase. For example, in lung adenocarcinomas, autocrine IL-6 has been identified as important activator responsible for the constitutive activation of STAT3 signaling [16]; furthermore, Maria reported that the absence of SOCS families molecules, including SOCS1, SOCS3, and SOCS5, promoted the propagation of the JAK/STAT3 signaling in the highly aggressive anaplastic thyroid cancer cells. Individual upregulation of SOCS1, SOCS3, and SOCS5 suppressed the activation of STAT3 signaling pathway activation resulting in alteration in the balance of proapoptotic and antiapoptotic molecules and sensitization to chemotherapeutic drugs in vitro [17]. These studies indicated that aberrant activation of JAK/STAT3 signaling pathway promoted the tumorigenesis and progression of cancer.

By being able to simultaneously inhibit a variety of target genes through binding to the 3′ untranslated region (3′UTR), aberrant expression of microRNAs (miRNAs) has been reported to be in multiple steps of cancer progression and metastasis [18], [19], [20], [21]. In the current study, our results reveal that miR-589-5p is dramatically elevated in HCC tissues and cells, which correlates with poor overall and relapse-free survival in HCC patients. Overexpression of miR-589-5p constitutively activates STAT3 signaling pathway via directly inhibiting multiple STAT3 signaling negative regulators, including SOCS2, SOCS5, PTPN1 and PTPN11. Furthermore, our results further demonstrate that upregulating of miR-589-5p enhances, while silencing miR-589-5p reduces the CSC-like characteristics and chemoresistance in vitro and in vivo. Importantly, recurrent gains contribute to miR-589-5p overexpression in HCC tissues. Collectively, our results demonstrate that miR-589-5p functions as an oncogenic miRNA in HCC, as well as uncover a novel mechanism for constitutive activation of STAT3 signaling in HCC.

Section snippets

Materials and methods

More methods used in this paper will be found in the Supplemental Materials.

miR-589-5p is upregulated in hepatocellular carcinoma and associated with poor survival

To discern the potential miRNA in the pathogenesis of HCC, the HCC miRNA sequencing dataset from The Cancer Genome Atlas (TCGA) was analyzed and the results revealed that miR-589-5p was dramatically elevated in HCC tissues compared with the adjacent normal tissues (Fig. 1A and B). Consistently, overexpression of miR-589-5p was observed in HCC miRNA dataset from E-GEOD-36918 and E-GEOD-31384 (Fig. S1 A and B). We further examined the miR-589-5p expression in our HCC tissues and found that

Discussion

It has been reported that STAT proteins were persistently phosphorylated on the tyrosine residue in variety of tumors, particularly STAT3, which further promoted proliferation, apoptosis, increased resistance to chemotherapeutic agents, cancer stem cell, leading to the progression and metastasis of cancer [28], [29]. STAT3 signaling has also been identified to promote the progression and metastasis via inducing cancer stem cell-like properties and chemotherapeutic resistance in different

Competing financial interests

No conflicts of interest were declared.

Acknowledgements

This study was supported by the Natural Science Foundation of China (No. 81300421, 81602701), the Natural Science Foundation of Guangdong Province (No. S2013040014343, 2014A030313090), the Science and Technology Projects Foundation of Guangdong Province (No. 2016A020215053), and the Doctoral Fund of Ministry of Education of China (No. 20130171120054).

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    Author contributions: Jiangting Long, Chunlin Jiang and Baoxian Liu contributed equally to this work.

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