Multiple defects in cell growth and differentiation may promote the transformation process of tumors[15], Gene mutation plays an important role in the pathogenesis of malignant tumor [16]. Up to now, many studies of oncogenes and tumor suppressor genes have helped to improve our understanding of the molecular pathophysiology of human carcinogenesis and tumor early detection, prediction of prognosis, and prevention.
In 1997, Dutch scholar Van Belzen et al. first reported NDRGl gene, and proved that the mRNA of this gene was up-regulated during the differentiation of colon epithelial cells and down-regulated in colorectal cancer. Since then, many scholars have carried out researches on this gene successively [17]. NDRG1 mainly exists in the cytoplasm, with some also located in the nucleus, cell membrane and intercellular link sites. More than seven phosphorylation sites have been reported on the NDRG1 protein [18]. Murray et al. showed that serum-and glucocorticoid-induced kinase 1, SGK1 phosphorylates NDRG1 at Thr346, Thr356, and Thr366 in the three tandem repeats, as well as at Thr328 and Ser330, conserved in all members of the NDRG family. SGK1 phosphorylation converts NDRG1 into a good substrate for serum- and glucocorticoid-induced kinase 3 (GSK3), and GSK3 phosphorylates Ser342, Ser352 and Ser362 of the NDRG1 tandem repeat [19]. In addition, NDRG1 in mast cells may be phosphorylated mainly on serine and threonine residues by protein kinase A (PKA), protein kinase C (PKC), and calmodulin kinase II [20]. These studies suggest more novel aspects of NDRG1, such as the enhancement of PKA/PKC phosphorylation by deleting three tandem repeats, and the effect of increased cell density on the reduction of NDRG1 phosphorylation. In hepatocellular carcinoma Hep3B cells, inhibition of HSP90 activity has been shown to decrease phosphorylation at Thr346, Thr356, and Thr366 on NDRG1[20]. In pancreatic cancer studies, phosphorylation of residues Ser330 and Thr346 in NDRG1 has been suggested to be critical in inhibiting the expression of the NF-KB signaling pathway and CXC chemokines [21].
The differences between mRNA and protein levels of NDRG1 in different tissues and organs suggest that the expression of NDRG1 is regulated by multiple levels of transcription and translation. NDRG1 gene has a certain response to various biological stimuli such as intracellular calcium ion level, nitric oxide, hypoxia, heavy metal ions, iron ions [4]. NDRG1 plays a role in normal cell growth, differentiation, and organ formation and is also highly responsive to hypoxia and oxidative stress, vitamin A, oncogenes (N-myc and c-myc), and tumor suppressor genes (p53 and PTEN) [22]. Studies have shown that the expression of NDRG1 in testis is mainly related to the level of androgen, and the expression of NDRG1 in prostate is mainly related to the development of reproductive organs[23, 24].Some studies have speculated that the abnormal expression of NDRG1 in cancer cells may be related to genetic or epigenetic regulation. Studies also have shown that the NDRG1 core promoter can inhibit NDRG1 by reducing its activity under the influence of the known oncogenes N-myc and c-myc [25]. Some scholars have found that lysine-specific demethylase 1 can cooperate with oncogene N-myc/MYCN to inhibit the expression of NDRG1 [26]. Furthermore, long non-coding RNA (lncRNA) NDRG1 overlapped transcript 1 can induce ubiquitination of NDRG1 protein under hypoxia conditions, leading to its proteasome degradation in breast cancer cells [27]. Therefore, it can be hypothesized that NDRG1 plays a multifactorial, specific, and pleiotropic role in cancer progression.
This study verified that NDRG1 mRNA was lowly expressed in gastric cancer tissues and high expression in paracancer tissues by RT-PCR experiment. Similarly, NDRG1 protein was still under expressed in gastric cancer tissues (high expression rate 34.62%, low expression rate 78.57%), while it was highly expressed in adjacent tissues (high expression rate 65.38%, low expression rate 21.43%). Therefore, it can be inferred that the expression of NDRG1 in gastric cancer tissues is lower than that in adjacent tissues at both mRNA and protein levels, which may be a tumor suppressor gene. At the same time, immunohistochemical results and clinicopathological data were analyzed, and it was found that NDRG1 was negatively correlated with the degree of tumor differentiation, the depth of tumor invasion and pTNM. Although the results showed that NDRG1 was not correlated with tumor size and lymph node metastasis, their statistical values were close to 0.05. We wondered whether the increase of sample size would make NDRG1 negatively correlated with tumor size and lymph node metastasis results. Therefore, it is concluded that NDRG1 may belong to the tumor suppressor gene and inhibit the occurrence and development of gastric cancer. This conclusion is in line with the findings of other scholars. For example, one study confirmed that overexpression of NDRG1 inhibits proliferation and invasion of gastric cancer cells, and induces G1 cell cycle arrest and early apoptosis [28]。
In recent years, there have been numerous reports on the mechanism of NDRG1 inhibiting the occurrence and development of malignant tumors. The first is the theory that NDRG1 inhibits stress fiber formation and cell collagen adhesion formation. Actin filaments (F-actin) are the primary mechanism of cell movement, often enhancing cancer cell migration and invasion [29], Stress fibers are actin bundles in the cell body, usually consisting of 10 to 30 F-actin[30]. A study by Sun et al. showed that overexpression of NDRG1 in prostate and colon cancer cells significantly reduced F-actin levels. In addition, a significant reduction in stress fibers was observed in cells overexpressing NDRG1 by immunofluorescence. Meanwhile, the expression of ROCK1 and Phosphorylation of MLC2 (pMLC2) were both significantly reduced in NDRG1 overexpressed cells[31]. pMLC2 is an important molecular motor for cell movement, and ROCK1 is the upstream molecule that induces pMLC2[32]. It can therefore be inferred that NDRG1 inhibits F-actin polymerization and stress fiber formation by inhibiting ROCK1/pMLC2 signaling pathway. Similarly, it has been shown that the FAK/paxillin signaling pathway enhances cell migration/invasion and regulates cytoskeleton formation of focal adhesion [33]. When activated, FAK induces phosphorylation of paxillin, which results in cell membrane prominence and enhancement and thus focal adhesion. It is noteworthy that the formation of focal adhesion affects cell migration and adhesion [34]. Studies have shown that overexpression of NDRG1 can significantly inhibit the phosphorylation of FAK at Tyr397, Tyr576/577 and Tyr925 in prostate and colon cancer cells, resulting in decreased cell adhesion. After the silencing of NDRG1 in cells, the phosphorylation of FAK at the above sites was significantly increased, resulting in increased cell adhesion [35]. It is well known that WNT/β-catenin signaling pathway can promote the proliferation, migration and invasion of cancer cells [36]. Studies have shown that NDRG1 can maintain the level of β-catenin in plasma membrane and inhibit the expression of cyclin D1, the downstream target protein of WNT/β-catenin signaling pathway, under the action of TGF-β. Meanwhile, NDRG1 silencing induces β-catenin nuclear translocation and increases cyclin D1 expression in colon and prostate cancer cells, Angiogenesis is one of the important factors promoting the occurrence and development of tumors. The growth of malignant tumors usually requires a large amount of oxygen and nutrients, and new blood vessels can further provide energy materials for tumor growth [37, 38]. In addition, angiogenic factors may also induce tumor invasion and metastasis [39]. Maruyama et al. found in mouse modeling experiments that overexpression of NDRG1 significantly reduced microvascular density and tumor-induced angiogenesis [40].
Because of the multiple functions shown by NDRG1 in cancer, several strategies have been proposed to target NDRG1. In osteosarcoma cells, NDRG1 plays an important role in lysosome function, and inhibition of NDRG1 enhances the antitumor effect of combretastatin A-4 and chloroquine combined therapy [41]. In radiation-resistant colorectal cancer cells, NDRG1 downregulation enhances DNA double-strand breaks and makes these cells radiation-sensitive [42]. These two experiments suggest that inhibition of expression of NDRG1 is considered as a strategy for targeted treatment of malignant tumors in which NDRG1 has a tumor-promoting effect. On the other hand, in malignant tumor types where NDRG1 has antitumor effects, upregulation of NDRG1 is considered a therapeutic strategy. In prostate cancer PC3 cells, the histone deacetylase inhibitor 2-valproic acid inhibits cell invasion by upregulation of NDRG1[43]. In PANC-1 cells of pancreatic cancer, another histone deacetylase inhibitor, trecomycin A, upregulates NDRG1 at both mRNA and protein levels and induces tumor cell type differentiation [44]. Some studies suggest that free radical nitric oxide regulates the expression of NDRG1 at mRNA and protein levels. It has been shown that NDRG1 is upregulated by the combination of intracellular nitric oxide with the chelated iron in the cell to form a dinitroso dimercaptan iron complex. Interestingly, nitric oxide does not require the expression of hypoxic-inducible factor 1α to induce upregulation of NDRG1 [45].
This study also has some shortcomings. They are concluded as follows. At first, the small sample size of the specimens prevented accurate assessment of the relationship between NDRG1 and tumor size and lymph node metastasis for the time being. In addition, due to the short time of specimen collection, no follow-up related to survival was conducted on the corresponding patients with specimen collection, making it impossible to accurately assess the effect of NDRG1 on the survival of patients with gastric cancer. Therefore, follow-up specimen collection and follow-up study are needed. A large number of cytological and zoological experiments are needed to determine whether NDRG1 can be used as a marker of tumor metastasis or even a target for drug therapy in the future.