A review on the role of NDRG1 in different cancers

NDRG1 is a member of the α/β hydrolase superfamily that resides in the cytoplasm and participates in the stress responses, hormone response, cell growth, and differentiation. Several studies have pointed to the importance of NDRG1 in the carcinogenesis. This gene has been found to be up-regulated in an array of cancer types such as bladder, esophageal squamous cell carcinoma, endometrial, lung and liver cancers, but being down-regulated in other types of cancers such as colorectal, gastric and ovarian cancers. The current study summarizes the evidence on the role of NDRG1 in the carcinogenic processes in different types of tissues.


Introduction
N-myc downstream regulated 1 (NDRG1) is encoded by a gene located on chromosome 8q24. 22, roughly 60 kb in size. This gene provides instructions for making a protein of 43 kDa (made up of 394 amino acids) that is extremely stable and is highly conserved among multicellular creatures. The mRNA for this gene is 3.0 kb in size [1] (Fig. 1). The NDRG1 gene is a part of the human NDRG family, which also includes the NDRG2, NDRG3, and NDRG4 genes. These genes are only 53-64% similar to one another [2]. As a member of the α/β hydrolase superfamily, this cytoplasmic protein takes part in the regulation of stress response, hormone response, cell growth, and differentiation. In addition, it has a crucial role in p53-mediated activation of caspase and apoptosis. Mutations in NDRG1 gene have been shown to cause Charcot-Marie-Tooth disease type 4D [3]. In addition to the cytoplasm, this protein can be found in the cell membrane and nucleus of the cells.
More recently, studies have pointed to the importance of NDRG1 in the carcinogenesis. This gene has been found to be up-regulated in an array of cancer types, but being down-regulated in other types of cancers. The current study summarizes the evidence on the role of NDRG1 in the carcinogenic processes in different types of tissues. We have categorized this paper into distinct subtitles based on the obtained evidence from cell lines, xenograft models of cancer and expression assays in clinical samples. 1 3 promoter. The HJURP/YAP1/NDRG1 axis can influence proliferation and chemosensitivity of TNBC cells [4].
Another study has re-analyzed the transcriptomic data from TNBC cybrids with a number of nuclear or mitochondrial donor cells. This study has shown up-regulation of 149 genes in the cybrids with mitochondria, among them being NDRG1. NDRG1 has been shown to be colocated with PVT1, and EXT1 on 8q24, a region that harbors amplification in breast cancer. NDRG1 has exhibited the most significant under-expression in the cybrids with benign mitochondria. NDRG1 silencing has led to reduction of proliferation of SUM-159 TNBC cells. Taken together, this study has demonstrated the role of mitochondria in facilitating over-expression of NDRG1 in TNBC [5].
Moreover, another study in breast cancer cells has shown that NDRG1 silencing leads to reduction of cell proliferation rate, abnormalities in lipid metabolism such as enhancement of incorporation of fatty acids into neutral lipids and lipid droplets. On the contrary, NDRG1 expression has diminished lipid droplet construction under nutrient supplied and starvation circumstances [6].
Besides, expression of NDRG1 can be affected by hormones. For instance, a cell line study has shown up-regulation of SGK1 and NDRG1 following treatment with progesterone. These observations have been accompanied by down-regulation of two miRNAs that target SGK1 3'UTR, namely miR-29a and miR-101-1. Authors have also demonstrated progesterone-mediated transcriptional and posttranscriptional control of SGK1 expression, resulting in over-expression of NDRG1 by a number of genes whose expressions are controlled by the transcription factor AP-1. NDRG1 can reduce activity of some kinases, thus reducing invasion and migratory potential of breast cancer cells [7].
In addition to hormones, expression of NDRG1 has been shown to be regulated by the long non-coding RNA (lncRNA) NDRG1 overlapping transcript 1 (NDRG1-OT1). Expression of this lncRNA is induced under hypoxic conditions. Notably, NDRG1-OT1 can inhibit expression of NDRG1 at both transcript and protein levels. NDRG1-OT1 enhances degradation of NDRG1 through ubiquitinmediated proteolysis. Moreover, NDRG1-OT1 can decrease NDRG1 promoter activity. Most notably, different segments of NDRG1-OT1 have been shown to exert opposite impacts on NDRG1. While the first quarter segment of this lncRNA has no impact on the promoter of NDRG1, the second and third quarter segments represses and improves NDRG1 promoter activity, respectively. Finally, the fourth quarter segment of this lncRNA decreases activity of this promoter through reducing KHSRP under hypoxia [8].
5-Aza-2′-deoxycytidine is another agent that induces expression of NDRG1. While NDRG1 is expressed in the MDA-MB-231, this gene is expressed in T47D cells only following treatment with 5-Aza-2′-deoxycytidine. This observation shows the impact of DNA methylation on NDRG1 expression. NDRG1 gene promoter has a high level of methylation in T47D cells in spite of low level of methylation of this promoter in MDA-MB-231 cells [9].
Furthermore, by modulating TLE2 and β-catenin expression, NDRG1 regulates Wnt pathway activation and EMT in esophageal cancer cells [10]. This suggests that NDRG1 in esophageal cancer cells plays a pro-oncogenic role through modulating tumor development (Fig. 2).
Additionally, K-Ras is critical for controlling in vitro NDRG1 protein level in pancreatic ductal adenocarcinoma (PDAC) cancer cells through ERK signaling [11]. Notably, NDRG1 expression reduces the amount and activity of HIF1 by inhibiting K-Ras downstream Akt/mTOR signaling. In particular, NDRG1 plays an especial part in controlling PDAC cancer metabolism. NDRG1 mediates downregulation of many glycolysis essential enzymes, including GLUT1, HK2, LDHA, and PDK1 [11]. Furthermore, it appears that NDRG1 inhibits glycolysis enzymes via decreasing the activity of HIF1, which may be the cause of this phenomenon.
Moreover, NDRG1 communicates with its target cells and other cells via signaling pathways and molecular motors. For instance, NDRG1 is associated with cancer metastasis via its function in Raf/MEK1/ERK signaling. Ras is bound to GTP upon activation of GFRs [12]. The downstream targets are activated when the active Ras:GTP phosphorylates Raf/ MEK1/ERK1/2. Smad4 is upregulated by NDRG1, which in turn blocks the pathway (Fig. 2).
Prostate cancer cell lines have been other sources for identification of the function of NDRG1. A chemical genetic screening method for identification of substrates for the oncogenic serine/threonine kinase PIM1 has led to identification of NDRG1 as an important substrate for this kinase in prostate cancer cells. PIM1 can phosphorylate pS330 of NDRG1, leading to reduction of stability NDRG1, its nuclear localization, and interaction with androgen receptor (AR), which consequently enhances cell migration and invasion [13]. Another study in prostate cancer has shown that N-cadherin induces expression of c-Jun and inhibits expression of NDRG1 to increase invasive properties and migratory potential of prostate cancer cells via affecting epithelial to mesenchymal transition (EMT). Moreover, c-Jun, AR, and DNMT1 establish a complex in the TPA response elements region of the NDRG1 promoter, which inhibits transcription of NDRG1 via induction of hypermethylation of DNA [14]. Two other studies have revealed a tumor suppressor role for NDRG1 in prostate cancer in association with the role of this protein in modulation of AR activity. First, NDRG1 has been shown to directly regulate AR signaling in this type of cancer [15]. Second, the impact of MLL5α on activation of AR/NDRG1 signaling has been found to result in the suppression of prostate cancer progression [16]. Table 1 summarizes the results of studies on the role of NDRG1 in different cancer cell lines.

Animal studies
The bulk of evidence from animal studies has indicated a tumor suppressor role for NDRG1 in colorectal, gastric, nasopharyngeal and renal cell cancers (Table 2). However, in osteosarcoma models, up-regulation of NDRG1 has been associated with higher rate of tumor growth [40]. Similarly, in non-small cell carcinoma, NDRG1 silencing has attenuated tumor growth and reduced angiogenesis [48].

Studies in clinical samples
NDRG1 has been suggested as a prognostic marker in patients with inflammatory breast cancer. Based on the results of univariate analyses, expression level of NDRG1, tumor grade, clinical stage and estrogen receptor (ER) status have been associated with overall and disease-free survival times. Patient with over-expression of NDRG1 have exhibited poor survival times. Particularly, those having overexpression of NDRG1 and ER-negative status have been shown to have worse outcome [59].
A meta-analysis of NDRG1 levels in numerous publicly available databases has shown correlation between NDRG1 up-regulation and expression of glycolytic and hypoxia-associated genes. Moreover, over-expression of NDRG1 has been associated with enhancement of metastasis and increase in patients' mortality [6]. Moreover, another study has revealed methylation of the NDRG1 promoter in about one third of primary breast cancer specimens. Moreover, methylation of promoter of this gene has been correlated with the TNM stage, metastasis, lymph node involvement, moderate and poor histological grade in these patients [9].
Proteomic analysis in human prostate cancer and benign prostatic hyperplasia samples have revealed association between over-expression of YWHAZ and NDRG1 and poor prognosis based on Gleason scores. In fact, YWHAZ and NDRG1 expression levels could define two groups of prostate cancer patients with high and intermediate risks of mortality. Based on the multivariable analyses, expression levels of these genes predict prognosis in an independent manner from Gleason scores [20].
Expression of NDRG1 has been found to be down-regulated in gastric cancer samples. Notably, its expression has been negatively correlated with the expression levels of DNMT1, DNMT3A and DNMT3B. Consistent with this finding, DNA methylation status of NDRG1 has been positively correlated to DNMT family. NDRG1 levels have been inversely correlated with invasion depth. However, levels of NDRG1 and DNMT1 have not been associated with prognosis of gastric cancer [26]. The latter finding is in contrast with another study in gastric cancer which has reported association between down-regulation of NDRG1 and poor clinical outcome [27].
Studies in other types of cancers have also revealed dysregulation of protein or mRNA levels of NDRG1 and   [58] associations between this abnormal expression pattern and malignant behavior of cancer as reflected in survival time of patients (Table 3).

Discussion
The role of NDRG1 has been vastly assessed in the contexts of breast, prostate, gastric and colorectal cancers. NDRG1 is an example of proteins with different roles in the carcinogenesis. These various and opposite effects can be at least partly explained by cell-or tissue-specific roles. However, experiments in a certain type of cancer have sometimes reported opposite effects for NDRG1. This is particularly true in breast and prostate cancers, two types of cancer that have been the focus of several independent studies. This discrepancy can be explained by differences in the stage, grade or other pathological features of cancer cells. Since tumor tissues are heterogeneous in terms of gene expression patterns, single cell gene expression profiling is needed to elaborate expression of NDRG1 in relation with pathological state. The lncRNA encoded by the same region (NDRG1-OT1) has been shown to effectively influence expression of NDRG1. Aryl hydrocarbon receptor and a number of miR-NAs mediate other mechanisms for regulation of expression NDRG1. Finally, methylation status of NDRG1 promoter has an established role in the regulation of its expression based on the results of studies in different tissues/ cell types.
Most importantly, the lncRNA encoded from NDRG1 locus has been shown to serve as a spong for miR-96-5p [38,60,61]. Since competing endogenous RNA function is an important mechanism for the regulation of expression of genes by lncRNAs, further studies should find other miR-NAs that are sponged by this lncRNA to unravel additional parts from the regulatory network of NDRG1.
Expression profiles of NDRG1 in clinical samples have revealed association between dysregulation of NDRG1 and poor clinical outcomes, demonstrating the prognostic impact of this protein in the context of cancer. However, the role of this protein in the early diagnosis of cancer, particularly based on expression assays in biofluids should be discovered in future.
Based on the importance of NDRG1 in the pathogenesis of cancer and induction or modulation of chemo-/    [57] radio-resistance in different types of cancers, future studies are required to unravel the exact role of this protein in the tissue-specific carcinogenic processes and develop specific therapies for each type of cancer. Taken together, the data presented above casts doubt on the previously supposed "anti-metastatic" role for NDRG1 and suggests a tissue-or cell-or stage-specific role for this protein in the carcinogenesis.