Research ArticlemiR-19 targeting of GSK3β mediates sulforaphane suppression of lung cancer stem cells☆
Introduction
Lung cancer is the leading cause of cancer mortality worldwide, accounting for about 13% of total cancer deaths [1], [2]. Despite significant improvement being made in cancer therapy, the 5-year relative survival rate of lung cancer still remains low [1]. Accumulating evidence has shown that cancer stem cells (CSCs), a rare subpopulation of cancer cells, possess the capacity of self-renewal and multipotent differentiation [3], [4]. CSCs express distinct and specific cell markers. For example, CD133, CD44 and aldehyde dehydrogenase (ALDH) are widely used to identify lung CSCs [5], [6]. It is well acknowledged that CSCs are responsible for the heterogeneity, metastasis and relapse of tumors [6]. Thus, the CSCs concept provides more effective intervention strategies in the prevention and treatment of cancers.
Several signal pathways are crucial in orchestrating CSCs activity. Dysregulation of these pathways has been implicated in the maintenance and function of CSCs. Aberrant Wnt/β-catenin signal pathway has been reported in several types of CSCs [7], [8], [9], [10]. The activation of Wnt/β-catenin pathway depends on the key regulator β-catenin, the intracellular level of whose is mainly modulated by glycogen synthase kinase-3β (GSK3β). In the absence of Wnt, the cytoplasmic β-catenin is phosphorylated by the destruction complex consisting of Axin, adenomatous polyposis coli (APC), casein kinase 1α (CK1α) and GSK3β and subsequently undergoes ubiquitin-proteasome degradation. Upon the Wnt ligand binding to the Frizzled receptor, β-catenin is dissociated from the destruction complex and accumulates in cytoplasm. The stabilized β-catenin then translocates into the nucleus where it forms a transcriptional complex with the T-cell factor (TCF)/lymphocyte enhancer factor (LEF), leading to the activation of downstream genes such as c-Myc, Cyclin D1, CD44, ALDH and others [11], [12], [13].
miRNAs are 21–25 nucleotides in length, small noncoding RNAs that regulate target genes by binding to the 3′-untranslated regions (3’UTRs), which leads to either degradation of mRNA or inhibition of protein translation [14]. Mounting evidence has indicated the pivotal role of miRNA in the development and progression of cancers by regulating CSCs activity [15], [16]. Some oncogenic miRNAs have been characterized, for example, the miR-17-92 cluster [17], [18], [19]. Among the miR-17-92 cluster, miR-19 is the key miRNA which exerts oncogenic function by targeting several critical tumor suppressor genes such as PTEN [20]. Previous studies revealed that miR-19 triggers the epithelial–mesenchymal transition (EMT) and is correlated with metastasis of lung cancer [21]. In gastric cancer, miR-19 promotes multidrug resistance (MDR) and regulates the self-renewal and proliferation of gastric CSCs [22], [23]. Moreover, a recent study has elucidated the role of miR-17-92 in lung cancer stem cells through regulation of Wnt signaling [24].
Numerous epidemiological studies have substantiated the efficient anticancer properties of dietary components in vegetables and fruits [25]. These bioactive and non/low-toxic phytochemicals have been proved to be promising candidates for cancer intervention [25], [26]. Sulforaphane (SFN), a major isothiocynate (ITC) abundant in broccoli or broccoli sprouts, has been shown to possess anticarcinogenic potential in vitro and in vivo [27]. Recently, it has been documented that SFN exhibits its anticancer property by targeting CSCs in various cancer types [28], [29], [30], [31]. In addition, by combining with other chemotherapeutic agents, SFN effectively abolishes CSCs characteristics [32]. To date, however, no studies have been conducted yet to examine the inhibitory effect of SFN on lung CSCs. Therefore, the present study aimed to investigate the regulatory role of the oncogenic miR-19 in lung CSCs and SFN modulation of lung CSCs.
Section snippets
Cell culture
Human lung cancer cell lines A549 and H1299 were purchased from Chinese Academy of Typical Culture Collection Cell Bank. Both cell lines were cultured in RPMI 1640 medium (Gibco, Carlsbad, CA, USA) containing 10% (v/v) fetal bovine serum and antibiotics (100-units/mL penicillin and 100-μg/mL streptomycin) in a humidified atmosphere of 5% CO2 at 37 °C.
Tumorsphere formation assay
One major feature of CSCs is their ability to form three-dimensional spheres in serum-free medium (SFM) culture condition. In SFM with some
miR-19a/19b is up-regulated in lung CSCs
CSCs are characterized by their capacity to form three-dimensional structures or spheres, and tumorsphere formation assay via serum-free medium (SFM) culturing is widely used in the isolation and enrichment of CSCs in vitro. These sphere-forming cells are considered to have CSCs characteristics like self-renewal and unlimited differentiation. In our study, turmorsphere formation assay was utilized to isolate and enrich lung CSCs from two human lung cancer cell lines A549 and H1299. Along with
Discussion
Emerging evidence has suggested that CSCs play essential role in the progress, metastasis and recurrence of human cancers, through their capacity for self-renewal, production of heterogeneous progeny, resistance to chemotherapy and unlimited proliferation. As important posttranscriptional regulators of gene expression, miRNAs are critically implicated in tumorigenic process. In the present study, we illustrated the pivotal role of miR-19 in regulating lung CSC traits and miR-19/GSK3β/β-catenin
Conflict of interest
The authors declare no conflict of interest.
Acknowledgements
This work was supported by grants from the National Natural Science Foundation of China (No. 81573139), the National Basic Research Program of China (973 Program) (No. 2013CB910303), the Science and Technology Planning Project of Guangdong Province of China (No. 2013B022000041), and by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
References (60)
- et al.
Lung cancer stem cell: fancy conceptual model of tumor biology or cornerstone of a forthcoming therapeutic breakthrough?
J Thorac Oncol
(2014) - et al.
Lung cancer stem cells: progress and prospects
Cancer Lett
(2013) - et al.
Post-translational glycoprotein modifications regulate colon cancer stem cells and colon adenoma progression in Apc(min/+) mice through altered Wnt receptor signaling
J Biol Chem
(2014) - et al.
Distinctive microRNA signature associated of neoplasms with the Wnt/beta-catenin signaling pathway
Cell Signal
(2013) Activator or inhibitor? GSK-3 as a new drug target
Biochem Pharmacol
(2013)- et al.
The miR-17-92 cluster expands multipotent hematopoietic progenitors whereas imbalanced expression of its individual oncogenic miRNAs promotes leukemia in mice
Blood
(2012) - et al.
MicroRNA-19a and -19b regulate cervical carcinoma cell proliferation and invasion by targeting CUL5
Cancer Lett
(2012) - et al.
Different micro-RNA expression profiles distinguish subtypes of neuroendocrine tumors of the lung: results of a profiling study
Mod Pathol
(2014) - et al.
MicroRNA-19 triggers epithelial-mesenchymal transition of lung cancer cells accompanied by growth inhibition
Lab Invest
(2015) - et al.
MicroRNA-19a/b regulates multidrug resistance in human gastric cancer cells by targeting PTEN
Biochem Biophys Res Commun
(2013)
Implications of cancer stem cell theory for cancer chemoprevention by natural dietary compounds
J Nutr Biochem
ALDH1 expression and the prognosis of lung cancer: a systematic review and meta-analysis
Heart Lung Circ
Wnt/beta-catenin signaling regulates cancer stem cells in lung cancer A549 cells
Biochem Biophys Res Commun
GSK-3 beta targets Cdc25A for ubiquitin-mediated proteolysis, and GSK-3 beta inactivation correlates with Cdc25A overproduction in human cancers
Cancer Cell
Inhibition of TBK1 attenuates radiation-induced epithelial-mesenchymal transition of A549 human lung cancer cells via activation of GSK-3beta and repression of ZEB1
Lab Invest
Cancer statistics, 2015
CA Cancer J Clin
Global cancer statistics, 2012
CA Cancer J Clin
Evidence for self-renewing lung cancer stem cells and their implications in tumor initiation, progression, and targeted therapy
Cancer Metastasis Rev
Cancer stem cells in lung cancer: evidence and controversies
Respirology
Nestin positively regulates the Wnt/beta-catenin pathway and the proliferation, survival and invasiveness of breast cancer stem cells
Breast Cancer Res
Cancer cells acquire a drug resistant, highly tumorigenic, cancer stem-like phenotype through modulation of the PI3K/Akt/beta-catenin/CBP pathway
Int J Cancer
PBK/TOPK enhances aggressive phenotype in prostate cancer via beta-catenin-TCF/LEF-mediated matrix metalloproteinases production and invasion
Oncotarget
Development of anticancer agents targeting the Wnt/beta-catenin signaling
Am J Cancer Res
Emerging role of microRNAs in cancer and cancer stem cells
J Cell Biochem
Targeting CSC-related miRNAs for cancer therapy by natural agents
Curr Drug Targets
The role of microRNAs in the regulation of cancer stem cells
Front Genet
miR-19 is a key oncogenic component of mir-17-92
Genes Dev
MiR-19b/20a/92a regulates the self-renewal and proliferation of gastric cancer stem cells
J Cell Sci
miR-17-92/p38alpha Dysregulation enhances Wnt signaling and selects Lgr6+ cancer stem-like cells during lung adenocarcinoma progression
Cancer Res
Targeting cancer stem cells with phytochemicals
Mol Interv
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Conflict of interest: The authors declare no conflict of interest.