MiRNAs and LncRNAs: Dual Roles in TGF-β Signaling-Regulated Metastasis in Lung Cancer

Lung cancer is one of the most malignant cancers around the world, with high morbidity and mortality. Metastasis is the leading cause of lung cancer deaths and treatment failure. MicroRNAs (miRNAs) and long non-coding RNAs (lncRNAs), two groups of small non-coding RNAs (nc-RNAs), are confirmed to be lung cancer oncogenes or suppressors. Transforming growth factor-β (TGF-β) critically regulates lung cancer metastasis. In this review, we summarize the dual roles of miRNAs and lncRNAs in TGF-β signaling-regulated lung cancer epithelial-mesenchymal transition (EMT), invasion, migration, stemness, and metastasis. In addition, lncRNAs, competing endogenous RNAs (ceRNAs), and circular RNAs (circRNAs) can act as miRNA sponges to suppress miRNAs, thereby mediating TGF-β signaling-regulated lung cancer invasion, migration, and metastasis. Through this review, we hope to cast light on the regulatory mechanisms of miRNAs and lncRNAs in TGF-β signaling-regulated lung cancer metastasis and provide new insights for lung cancer treatment.


Introduction
Lung cancer is the most common cancer based on current diagnoses and the leading cause of cancer-related deaths globally [1]. According to the data from the GLOBOCAN database, it is estimated that 2.09 million new cases (11.6% of total cancer cases) and a mortality of 1.76 million (18.4% of total cancer-related deaths) of lung cancer were found worldwide in 2018 [2]. Lung cancer can be divided into small cell lung cancer (SCLC) (15%) and non-small cell lung cancer (NSCLC) (85%). NSCLC contains three major subtypes: lung adenocarcinoma (40%), lung squamous cell carcinoma (25-30%), and large cell carcinoma (10-15%) [3,4]. Lung cancer development is mostly correlated with genetic mutations and environmental interactions, such as cigarette smoking, radiation exposure, and toxic substances [5]. Presently, metastasis and recurrence are the main causes of death for lung cancer patients [6]. Statistics show that approximately two-thirds of patients have metastatic lesions when they are first diagnosed with lung cancer [7]. Although there are many therapies (surgical resection, chemotherapy, radiotherapy, etc.) to treat lung cancer, the overall five-year survival rate of lung cancer patients is still as low as 17.7% [8], and the overall five-year survival rate of patients with distant organ metastasis is 4.5% [9]. Therefore, it is necessary to explore the metastatic mechanisms of lung cancer, as finding new strategies to diagnose and treat lung cancer is of great significance.
MicroRNAs (miRNAs) and long non-coding RNAs (lncRNAs) are small non-coding RNAs (nc-RNAs) without the potential to encode proteins [10]. Nevertheless, these RNAs can regulate gene transcription and translation by interacting with the 3 untranslated region (3 UTR) of messenger Figure 1. TGF-β signaling regulates lung cancer cell invasion and migration. TGF-β can mediate EMT via Smad pathways and non-Smad pathways. TGF-β can increase Snail, Slug, and ZEB2 expression to decrease E-cadherin. It can also bind to TGFBR2 and then phosphorylate Par6 to recruit Smad ubiquitin regulatory factor 1 (SMURF1). SMURF1 can degrade RhoA. These can reduce cell-cell adhesion. TGF-β1 can activate PI3K/AKT/NF-κB to increase β1 integrin expression. TGF-β/Smad3 signal can increase α5β1 integrin expression by down-regulating zyxin. These can enhance cell-matrix adhesion. TGF-β can increase phosphorylated focal adhesion kinase (pFAK), Src, cortactin, and ERK in response to β3 integrin, resulting in forming a mature invadopodia.
TGF-β can induce Snail, Slug, and ZEB2 expression to decrease E-cadherin, resulting in breaking adherens junctions [40]. TGF-β can bind to TGFBR2 and then phosphorylate Par6 to recruit SMURF1. Then, SMURF1 can degrade RhoA to increase the dissolution of junctional complexes and thereby reduce cell-cell adhesions [41]. In contrast to cell-cell adhesions, cell-matrix adhesions are always enhanced in lung cancers ( Figure 1). In A549 cells, TGF-β1 can activate the PI3K/AKT/NF-κB signaling pathway to increase β1 integrin expression, which may facilitate integrin-mediated cell migration [42]. TGF-β/Smad3 signal can increase α5β1 integrin expression by down-regulating zyxin. Up-regulated α5β1 integrin can enhance A549 cell adhesion to the fibronectin, as well as increase migration [43].

TGF-β Signaling Regulates Lung Cancer's Distant Metastasis
Lung tumor cell intravasation is involved in migrating to and then passing through the vessel wall. Tumor cells not only secrete TGF-β1 but also bind to Notch ligands on endothelial cells (ECs) by using Notch receptors, thus facilitating their transmigration through the endothelial tight junctions [47]. TGF-β can increase β3 integrin expression to enhance H157 cell adhesion to Figure 1. TGF-β signaling regulates lung cancer cell invasion and migration. TGF-β can mediate EMT via Smad pathways and non-Smad pathways. TGF-β can increase Snail, Slug, and ZEB2 expression to decrease E-cadherin. It can also bind to TGFBR2 and then phosphorylate Par6 to recruit Smad ubiquitin regulatory factor 1 (SMURF1). SMURF1 can degrade RhoA. These can reduce cell-cell adhesion. TGF-β1 can activate PI3K/AKT/NF-κB to increase β1 integrin expression. TGF-β/Smad3 signal can increase α5β1 integrin expression by down-regulating zyxin. These can enhance cell-matrix adhesion. TGF-β can increase phosphorylated focal adhesion kinase (pFAK), Src, cortactin, and ERK in response to β3 integrin, resulting in forming a mature invadopodia.
TGF-β can induce Snail, Slug, and ZEB2 expression to decrease E-cadherin, resulting in breaking adherens junctions [40]. TGF-β can bind to TGFBR2 and then phosphorylate Par6 to recruit SMURF1. Then, SMURF1 can degrade RhoA to increase the dissolution of junctional complexes and thereby reduce cell-cell adhesions [41]. In contrast to cell-cell adhesions, cell-matrix adhesions are always enhanced in lung cancers ( Figure 1). In A549 cells, TGF-β1 can activate the PI3K/AKT/NF-κB signaling pathway to increase β1 integrin expression, which may facilitate integrin-mediated cell migration [42]. TGF-β/Smad3 signal can increase α5β1 integrin expression by down-regulating zyxin. Up-regulated α5β1 integrin can enhance A549 cell adhesion to the fibronectin, as well as increase migration [43].

TGF-β Signaling Regulates Lung Cancer's Distant Metastasis
Lung tumor cell intravasation is involved in migrating to and then passing through the vessel wall. Tumor cells not only secrete TGF-β1 but also bind to Notch ligands on endothelial cells (ECs) by using Notch receptors, thus facilitating their transmigration through the endothelial tight junctions [47]. TGF-β can increase β3 integrin expression to enhance H157 cell adhesion to lymphatic endothelial cells (LECs), subsequently promoting cell transmigration across LEC monolayers [48]. The consequences of successful intravasation are indicated by the appearance of circulating tumor cells (CTCs) in circulation.
CTCs are usually arrested in narrow capillaries, where they can be trapped because of the size restriction and then extravasate [53]. In A549 cells, the TGF-β/Smad3 signaling pathway can induce a DOCK4 expression to activate Rac1. Then, Rac1 activation can extend protrusions to promote the extravasation of A549 cells ( Figure 2) into distant organ sites [54]. Lung cancer cells are most likely to form metastases in the brain, bones, and lymphatic nodes [55]. In H460 cells, TGF-β can increase Figure 2. TGF-β signaling regulates lung cancer's distant metastasis. TGF-β can increase β3 integrin expression to enhance cancer cell intravasation. TGF-β can increase Tregs via Cox-2/PGE2 to suppress the proliferation of CD4+ T cells, thus driving lung CTCs to escape from immune attacks. TGF-β can induce miR-183 to down-regulate MICA and MICB expression, which can help cancer cells evade immune detection. TGF-β1/Smad2 can up-regulate PD-L1. PD-L1 can bind to PD-1 to suppress anti-tumor immunity. TGF-β/Smad3 can induce dedicator of cytokinesis 4 (DOCK4) expression to activate Rac1. Then, Rac1 activation can promote cancer cell extravasation. TGF-β/Smad2/3 is required for the combination of histone deacetylase 4 (HDAC4), paired-like homeodomain 1 (PITX1), and roundabout, axon guidance receptor, and homolog 1 (ROBO1), which can promote the bone colonization of lung cancer cells. TGF-β can increase transcription factor 4 (TCF4), protein kinase D 3 (PRKD3), and SUSD5 levels, which can also enhance bone colonization.
CTCs are usually arrested in narrow capillaries, where they can be trapped because of the size restriction and then extravasate [53]. In A549 cells, the TGF-β/Smad3 signaling pathway can induce a DOCK4 expression to activate Rac1. Then, Rac1 activation can extend protrusions to promote the extravasation of A549 cells ( Figure 2) into distant organ sites [54]. Lung cancer cells are most likely to form metastases in the brain, bones, and lymphatic nodes [55]. In H460 cells, TGF-β can increase the levels of TCF4, PRKD3, and SUSD5. The combination of these triple factors can enhance osteolytic lesions that can increase bone colonization [56]. In A549 cells, TGF-β/Smad2/3 signaling is required for the combination of HDAC4, PITX1 and ROBO1. This combination can promote osteolytic lesions, thereby promoting bone colonization ( Figure 2) of A549 cells [57]. MiR-93 is up-regulated in NSCLC tissues [58]. The neural precursor cell expressed developmentally downregulated gene 4-like (NEDD4L) was predicted to be its target by miRDB and TargetScan. MiR-93 can bind to the mRNA 3 UTR of the NEDD4L and then down-regulate its expression. Down-regulated NEDD4L can increase Smad2 phosphorylation to enhance TGF-β signal transduction, finally promoting TGF-β-induced EMT in A549 and H1650 cells [59]. In addition, increased miR-93 can predict poor outcomes among NSCLC patients [60].

MiR-128-3p
MiR-128-3p is highly expressed in NSCLC tissues compared to adjacent normal tissues. It can inhibit SMURF2 and the catalytic subunit of protein phosphatase 1 (cPP1) expressions by targeting their mRNA 3 UTRs, eventually activating the TGF-β signaling pathways to promote EMT, metastasis, and chemoresistance. Further, high miR-128-3p expression can indicate poor NSCLC patient survival [61].

MiR-134 and miR-487b
MiR-134 and miR-487b are both located on chromosome 14q32, and both of them can be activated by TGF-β1 during EMT progression in lung adenocarcinoma. Moreover, over-expressed miR-134 and miR-487b can promote TGF-β1-induced EMT by suppressing membrane-associated guanylate kinase, WW and PDZ domain-containing protein 2 (MAGI2) (a multidomain scaffolding protein that can repress Smad3-induced transcriptional activity by interacting with Smad3 [64]) in A549 cells [65].

MiR-1246
MiR-1246 expression has been shown to be markedly increased in NSCLC and SCLC patients' serum by quantitative real-time polymerase chain reaction (qRT-PCR). It can decrease E-cadherin, as well as increase vimentin and TGF-β, contributing to promoting EMT in A549 cells [68].

MiR-9-5p
MiR-9-5p levels are higher in NSCLC tissues and cell lines compared to normal ones. It has been revealed that decreased receptor effects can abrogate TGF-β suppressive functions, thereby showing its oncogenic roles in NSCLC [69,70]. MiR-9-5p can bind to TGFBR2 mRNA and then attenuate its expression, subsequently facilitating A549 cells invasion and migration [71].

MiR-181b-5p
Cancer stem cells (CSCs) possess self-renewal properties and have been demonstrated to be a small population of cells within lung cancer tissues, which can promote lung tumor metastasis [72]. MiR-181b-5p can be obviously up-regulated by TGF-β1 in lung CSCs and A549 cells. It can suppress E-cadherin expression, thereby promoting EMT and eventually increasing invasion and metastasis in lung CSCs and A549 cells in vitro and in vivo. Moreover, up-regulated miR-181b-5p can predict poor outcomes in NSCLC patients [73].

MiR-23a
MiR-23a is up-regulated in NSCLC cell lines, and can also be up-regulated by the TGF-β1/Smad2/3 signaling pathway. MiR-23a can target the 3' UTR of CDH1 (a gene that can encode E-cadherin) and then suppress E-cadherin expression, eventually enhancing TGF-β1-induced EMT in A549 cells. Further, miR-23a can also cause resistance to gefitinib [74]. Accumulating studies have revealed that lung cancer cells can secrete exosomes into the extracellular environment to affect lung tumor cell invasion, migration, and metastasis. In this process, exosomes can carry several materials, such as DNA, nc-RNAs, and proteins [75]. MiR-23a and β-catenin can be increased by TGF-β1 in A549 cells-derived exosomes, while E-cadherin is decreased ( Figure 3). In addition, treatment with these exosomes can induce EMT in A549 cells [76]. MiR-9-5p levels are higher in NSCLC tissues and cell lines compared to normal ones. It has been revealed that decreased receptor effects can abrogate TGF-β suppressive functions, thereby showing its oncogenic roles in NSCLC [69,70]. MiR-9-5p can bind to TGFBR2 mRNA and then attenuate its expression, subsequently facilitating A549 cells invasion and migration [71].
Cancer stem cells (CSCs) possess self-renewal properties and have been demonstrated to be a small population of cells within lung cancer tissues, which can promote lung tumor metastasis [72]. MiR-181b-5p can be obviously up-regulated by TGF-β1 in lung CSCs and A549 cells. It can suppress E-cadherin expression, thereby promoting EMT and eventually increasing invasion and metastasis in lung CSCs and A549 cells in vitro and in vivo. Moreover, up-regulated miR-181b-5p can predict poor outcomes in NSCLC patients [73].

MiR-23a
MiR-23a is up-regulated in NSCLC cell lines, and can also be up-regulated by the TGF-β1/Smad2/3 signaling pathway. MiR-23a can target the 3' UTR of CDH1 (a gene that can encode E-cadherin) and then suppress E-cadherin expression, eventually enhancing TGF-β1-induced EMT in A549 cells. Further, miR-23a can also cause resistance to gefitinib [74]. Accumulating studies have revealed that lung cancer cells can secrete exosomes into the extracellular environment to affect lung tumor cell invasion, migration, and metastasis. In this process, exosomes can carry several materials, suc  In this process, TGF-β can decrease E-cadherin and increase β-catenin, miR-23a and lnc-MMP2-2. Decreased E-cadherin and increased β-catenin can then promote EMT. Increased miR-23a can decrease E-cadherin to enhance EMT. Up-regulated lnc-MMP2-2 can inhibit E-cadherin, as well as increase N-cadherin and vimentin to promote invasion and migration; it can also increase vascular permeability by repressing occludin and Zo-1. Moreover, lnc-MMP2-2 can augment MMP2 to accelerate metastasis.

MiR-132
In NSCLC tissues with lymphatic metastasis, the expression of miR-132 is found to be obviously lower than that in adjacent normal tissues. MiR-132 can suppress the TGF-β1/Smad2 signaling pathway and, moreover, reduce N-cadherin, ZEB1, Snail, and vimentin, leading to a repression of EMT, invasion, and migration in A549 cells [77].

MiR-203 and miR-145
MiR-203 and miR-145 are reduced in NSCLC tissues and cell lines. Smad3 was predicted to be their target by PicTar and miRanda. Both miR-203 and miR-145 can interact with the 3 UTR of Smad3 mRNA to decrease its levels, ultimately inhibiting TGF-β1-induced EMT and invasion in A549 and 95C cells [78].

MiR-16
MiR-16 is decreased in human NSCLC tissues compared to adjacent normal tissues. Autophagy is a complex biological process associated with the lysosomal degradation of cellular proteins and other cytoplasmic contents; it can also suppress NSCLC metastasis [92,93]. MiR-16 mimics were transfected into A549 cells to increase their expression levels. Up-regulated miR-16 can activate autophagy to inhibit TGF-β1-induced EMT [94].

MiR-3607-3p
MiR-3607-3p expression is lower in NSCLC tissues than that in adjacent normal lung tissues. It can bind to the 3 UTR of both TGFBR1 and cyclin E2 (CCNE2) mRNAs and then repress their expressions, thereby inhibiting NSCLC invasion and migration [99].

MiR-136
MiR-136 has been found to be over-expressed in NSCLC tissues compared to adjacent normal lung tissues [101]. In A549 and Calu-3 cells, miR-136 can reduce cell invasion and migration by reducing Smad2 and Smad3 expression, which may be involved in attenuating the TGF-β signaling pathway [102].

MiR-133a
MiR-133a is also down-regulated in lung adenocarcinoma cells. MiR-133a can abrogate AKT signaling by reducing the TGF-β/TGFBR1 signal, contributing to suppressing A549 cell invasion and migration. In addition, low miR-133a expression can indicate a poor overall survival for lung cancer patients [103].

MiR-143
MiR-143 is obviously reduced in NSCLC tissues compared to normal lung tissues. It can be stimulated by the TGF-β/Smads signaling pathway. MiR-143 can inhibit Smad3, CD44, and K-Ras expression, eventually repressing cell invasion and migration in A549 cells [104].

MiR-138
MiR-138 can be down-regulated by TGF-β1 in LC006 and LC021 cells. It can inhibit CSC formation by suppressing TGF-β1-induced EMT in primary lung cancer cells [106].

LncRNA HCP5
LncRNA histocompatibility leukocyte antigen complex P5 (HCP5) is highly expressed in lung adenocarcinoma cell lines, especially in current smokers and patients with mutated EGFR and KRAS. HCP5 can be induced by the TGF-β/Smad3 signaling pathway. In addition, it can augment Snail and Slug expression by attenuating miR-203, which can promote TGF-β/Smad3 signaling-induced EMT and invasion. Furthermore, HCP5 over-expression can indicate a poor prognosis for lung adenocarcinoma patients [107].

LncRNA NORAD
LncRNA NORAD expression is higher in NSCLC tissues than in adjacent normal tissues [108]. NORAD can stimulate Smad3 and importin β1 complex formation and then promote the TGF-β-induced translocation of the Smad2/3 complexes into the cell nucleus. NORAD ultimately increases EMT-related gene (SNAI1 and FN1, which can encode Snail and fibronectin, respectively) expression to enhance TGF-β-induced EMT [18].

LncRNA XIST
LncRNA X inactivate-specific transcript (XIST) is up-regulated in NSCLC tissues. On the one hand, XIST can bind to miR-137 and decrease its expression, thereby suppressing miR-137 s inhibitory roles in TGF-β1-induced EMT [20]. On the other hand, it can act as a kind of miRNA sponge to reduce miR-367 and miR-141 expression, thereby increasing ZEB2 [19]. Through these two mechanisms, XIST can facilitate TGF-β1-induced EMT in A549 cells [19,20].

LncRNA linc00673
LncRNA linc00673 is up-regulated in NSCLC tissues and cell lines. It can also be elevated by TGF-β. MiR-150-5p can target ZEB1 mRNA 3 UTR and attenuate its expression. Linc00673 can act as a competing endogenous RNA (ceRNA) that can suppress the effect of miR-150-5p on ZEB1, ultimately potentiating TGF-β-induced EMT, invasion, and migration in A549 and H1975 cells. Furthermore, up-regulated linc00673 can indicate poor survival for NSCLC patients [109].

LncRNA MEG3
LncRNA MEG3 can be overexpressed by TGF-β during EMT progression. In A549 and LC-2/ad cells, MEG3 shRNA can silence its expression, and MEG3 knockdown can abrogate TGF-β-induced actin stress fiber formation, which can inhibit EMT. MEG3 cDNA can increase its levels, and MEG3 over-expression can not only decrease the expression of genes CDH1, miR-200a, and miR-200c but can also increase the expression of genes ZEB1 and ZEB2 through the recruitment of JARID2 and the enhancer of zeste homologue 2 (EZH2) [110].

LncRNA MEG8
LncRNA MEG8 can also be up-regulated by TGF-β during EMT progression. MEG8 can interact with EZH2 and can subsequently down-regulate CDH1 expression. At the same time, MEG8 can up-regulate SNAI1 and SNAI2 (a gene that can encode Slug) expression by repressing the miR-34a and miR-203 genes in A549 and LC-2/ad cells. It has also been revealed that neither MEG8 nor MEG3 can exclusively lead to EMT without TGF-β. However, the co-expression of MEG8 and MEG3 can induce EMT in the absence of TGF-β [111].

Lnc-MMP2-2
Lnc-MMP2-2 is over-expressed in lung cancer tissues, and it can be obviously increased in the exosomes derived from A549 cells by TGF-β (Figure 3). On the one hand, exosomal lnc-MMP2-2 can increase N-cadherin and vimentin and decrease E-cadherin to facilitate invasion and migration. On the other hand, it can inhibit occludin and zonula occludens-1 (Zo-1) to impair vascular tight junctions, thus increasing vascular permeability. Exosomal lnc-MMP2-2 can also increase MMP2, which can accelerate metastasis [116].

LncRNA AWPPH
LncRNA AWPPH expression was estimated to be higher in the blood of lung cancer patients than in normal people by qRT-PCR. AWPPH can stimulate TGF-β1 protein expression to promote NSCLC migration and invasion. Moreover, high AWPPH can indicate the postoperative distant recurrence of NSCLC patients [117].

LncRNA CASC11
LncRNA CASC11 has been shown by qRT-PCR to be significantly increased in the plasma of SCLC patients. CASC11 can promote SCLC stemness by up-regulating TGF-β1, eventually increasing the number of lung CSCs. Further, CASC11 up-regulation can reveal the poor prognosis of SCLC patients [118].

LncRNA linc01186
LncRNA linc01186 is decreased in NSCLC tissues. It can be repressed by the TGF-β1/Smad3 signaling pathway. Linc01186 can inhibit cell invasion and migration by repressing EMT [120].

LncRNA ANCR
LncRNA ANCR has been shown by qRT-PCR to be markedly decreased in the plasma of NSCLC patients. Moreover, it can decrease TGF-β1 expression and thus inhibit NSCLC migration and invasion. Low ANCR expression can reveal the shorter overall survival time of patients than that of patients with high ANCR expression [122].

MiRNA Sponges Participate in TGF-β Signaling-Regulated Lung Cancer Metastasis
LncRNAs, ceRNAs, and circular RNAs (circRNAs) have been regarded as miRNA sponges to inhibit miRNAs in lung cancer. LncRNA has MREs, and it can influence the target gene downstream of the miRNAs by repressing them [124]. CeRNAs can regulate gene expression by competitively binding to miRNA through MREs, thereby suppressing the interactions between miRNA and its targeted mRNA. CeRNA is also essential for miRNA and lncRNA regulatory networks [125]. CircRNAs, belonging to endogenous nc-RNAs, possess closed-loop structures without a 5' end cap and a 3' end poly tail structure. CircRNA has several miRNA-binding sites to adsorb miRNA, subsequently counteracting miRNA's effects on its target gene expression [126]. Additionally, circRNAs can also work as ceRNAs in lung cancer. CeRNAs and circRNAs can regulate lung cancer migration, invasion, and metastasis [127,128].

CeRNA HMGA2
CeRNA HMGA2 is up-regulated in metastatic lung adenocarcinoma. HMGA2 can increase TGFBR3 expression by repressing miRNA let-7, which can also enhance the TGF-β signaling pathway, ultimately promoting invasion and metastasis in NSCLC [130].

Targeted Therapy
Although traditional lung cancer therapies, such as surgical resection, chemotherapy, and radiotherapy, have made great progress, they still face some limitations. For example, drug resistance and lung cancer distant metastasis can usually influence the therapeutic effects of a treatment. As a result, it is necessary to develop new strategies. MiRNAs and lncRNAs participate in substantial lung cancer progression, and we may be able to use them as treatment targets.

Curcumin
Curcumin, isolated from Curcuma longa, possesses anti-cancer effects and chemopreventive properties. Curcumin can significantly inhibit several miRNA-associated pathways, such as the MAPK, TGF-β, and Wnt signaling pathways, contributing to suppressing invasion and migration in A549 cells [133].

Carboplatin
Carboplatin can interact with ifosfamide and paclitaxel and possesses fewer non-hematologic toxicities than cisplatin (DDP). Carboplatin is a chemotherapeutic drug widely used to treat SCLC and NSCLC in order to improve overall survival [134]. Carboplatin can suppress miR-21 to increase Smad7 expression. Then, up-regulated Smad7 can inhibit the TGF-β/Smads signaling pathways, thereby suppressing TGF-β-mediated invasion in A549 cells [135].

DDP
DDP is also extensively used to deal with lung cancer. MiR-181b is decreased in NSCLC cells. It can target TGFBR1 mRNA 3 UTR and repress its expression, thus abrogating the TGF-β signal and eventually increasing DDP chemosensitivity in NSCLC cells [136]. MiR-17, miR-20a, and miR-20b are all down-regulated in A549/DDP cells (DDP-resistant A549 cells). They can attenuate the TGF-β signal pathway by inhibiting TGFBR1, thereby reducing migration and increasing DDP sensitivity in A549/DDP cells [137].

DAC
Decitabine (DAC) is a deoxyribonucleotide and can abrogate DNA methylation. Functionally, DAC promotes demethylation of the miR-200 promoter to increase the levels of miR-200c and miR-200a, which can repress ZEB1 and ZEB2 expression, leading to decreasing TGF-β1-induced EMT and metastasis in PC9 cells [138].

Discussion and Conclusions
Lung cancer is the most common cancer in men and the second most common in women after breast cancer. The low overall survival rate of lung cancer patients is usually correlated with distant organ metastasis. MiRNAs and lncRNAs are demonstrated to be oncogenes or suppressors by targeting mRNA 3 UTR in lung cancer development. Currently, miRNAs and lncRNAs are widely used to explore lung cancer's metastatic mechanisms, and these two nc-RNAs are increasingly used as biomarkers to predict the outcomes of lung cancer patients. TGF-β is involved in many cancers, and TGF-β signaling pathways have also been extensively confirmed to play pivotal roles in lung cancer metastasis. Interestingly, we found that Carboplatin, the first-line therapy used to treat metastatic lung cancer, suppresses TGF-β signaling-regulated lung cancer metastasis by repressing miR-21 [135]. This result suggests a potential treatment strategy involving miRNAs and TGF-β signaling for lung cancer metastasis. Considering the widespread attention attracted by the rational design of miRNA/lncRNA-targeted drugs against lung cancer, we have summarized and discussed the relationships between these two nc-RNAs and TGF-β signaling pathways in lung cancer metastasis. We hope that this review will provide new ideas for more clinical therapies like Carboplatin, thereby allowing us to treat lung cancer more effectively.
MiRNAs and lncRNAs can be detected in biological fluids through a biopsy of liquids, such as whole blood (lncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) [141] and miRNA-182-5p [142]), serum (lncRNA linc00342 [143], MEG3 [144], miR-96 [145] and miR-30a-5p [146]), plasma (lncRNA linc00152 [147], MALAT1 [148], miR-145 [149], miR-30b [150], miR-25 [151] and miR-425-3p [152]), sputum (lncRNA HOTAIR, H19 [153], miR-21 and miR-210 [154]), and pleural effusion (MALAT1 [155] and miR-200 family [156]) in lung cancer. Furthermore, miRNAs can also be found in bronchoalveolar lavage fluid (miR-126 and miR-Let-7a [157]). Both lncRNAs and miRNAs are usually dysregulated in lung cancer and are closely related to cancer metastasis. For instance, miR-25 is up-regulated in NSCLC tissues and cell lines and can repress large tumor suppressor homology 2 (LATS2) and then increase the yes-associated protein (YAP), ultimately contributing to promoting A549 cells migration and invasion [158]. Moreover, high plasma miR-25 levels can indicate a poor prognosis for NSCLC patients [151]. MiR-96 is elevated in the serum-derived exosomes of lung cancer patients. MiR-96 can suppress LIM-domain only protein 7 (LMO7) expression, thereby promoting A549 cell proliferation, migration, and DDP resistance. In addition, increasing serum exosomal miR-96 can predict lung cancer lymph node metastasis [145]. MiR-21-5p is highly expressed in NSCLC tissues. It can decrease E-cadherin and increase MMP9 and vimentin, which can facilitate A549 cell invasion and migration [159]. Further, high plasma miR-21 can reveal the distant metastasis of NSCLC patients [160]. The expression of MALAT1 is significantly high in NSCLC tissues and cell lines [161]. MALAT1 can enhance A549 cell invasion and migration by repressing miR-200a-3p [162]. High serum exosomal MALAT1 could be an indicator for lymphatic metastasis in NSCLC patients [163]. Linc00152 is up-regulated in lung adenocarcinoma tissues and can enhance H1299 and H1975 cell migration and invasion [164]. Additionally, increased plasma linc00152 levels can indicate recurrence in NSCLC patients [147]. Linc00342 is up-regulated in NSCLC tissues and cell lines. Linc00342 can increase cell invasion and migration by inhibiting miR-203a-3p in NCI-H1299 and NCI-H460 cells [165], and high serum linc00342 can act as a better biomarker to diagnose NSCLC than CYFRA 21-1 (a sensitive factor that can diagnose lung cancer squamous cell carcinoma [166]) [143]. In addition to circulating miRNAs and lncRNAs, the existence of lung cancer CTCs can also predict a poor outcome for lung cancer patients. For example, the presence of CTCs at six months after surgery can increase the risk of metastasis and the short relapse-free survival of lung cancer patients [167]. It has been shown that molecular characterizations (proteins, RNAs, etc.) are closely related to lung cancer detection, prognosis, and therapeutic efficacy [168]. Therefore, miRNA, as a type of molecular characterization, might provide new ideas for the diagnosis of lung cancer metastasis, prognostic prediction, and therapeutic selection in lung cancer patients.
Due to the multiple functions of miRNAs and lncRNAs in lung cancer metastasis, these RNAs have been regarded as promising therapeutic targets. Several traditional treatments have been discovered to target miRNAs and lncRNAs. Low-dose radiation can increase miR-30a and miR-30b expression. MiR-30a and miR-30b can inhibit plasminogen activator inhibitor-1 (PAI-1) expression and then reduce EMT and migration in NCI-H460 cells exposed to high-dose radiation. Additionally, miR-30a and miR-30b also promoted sensitivity to radiotherapy in a mouse model [169]. In Lewis lung cancer cells, radiation can attenuate HOTAIR expression to reduce the nuclear localization of β-catenin, thereby inhibiting tumor cell proliferation [170]. The effective chemotherapeutic drug, paclitaxel (PTX), can increase MEG3, thus promoting A549 cell apoptosis by increasing p53 expression. Further, over-expressed MEG3 can strengthen PTX's anti-tumor activity in A549 cells [171]. DDP can increase miR-425-3p in cells and NSCLC cell-derived exosomes. Exosomal miR-425-3p can activate autophagy by inhibiting AKT1, ultimately repressing DDP sensitivity in A549 cells [172]. MiRNAs can also exert therapeutic effects. MiR-200c can be delivered to lung tumor cells through nano vehicles. Nano miR-200c can increase the expressions of the tumor suppressors miR-29b and miR-1247 in metastatic and non-metastatic lung cancer cells. It can also suppress lung tumor cell migration and invasion [173]. Tumor-derived exosomes can selectively deliver anticancer therapies to lung cancer sites due to their intrinsic organotropic tumor-homing properties [174][175][176]. Functionalization of exosome-mimetic nanosystems (EMNs) with ITGα6β4 (F-EMNs) have similar structures and capabilities against tumor-derived exosomes and can escape endolysosomal degradation. MiR-145 can repress N-cadherin expression and then inhibit A549 cell migration and invasion [177]. The injection of F-EMNs loaded with therapeutic oligonucleotide miR-145 can effectively reach lung tumor sites. Despite the nonnegligible safety concerns caused by their low systemic toxicities, this method remains a promising approach to target lung cancer cells during the targeted therapy process [178]. It has been indicated that nanomedicine inhalation can also be used to treat lung cancer [179]. One study found that siRNA combined with PTX inhalation via nanostructured lipid carriers can significantly decrease lung tumor volumes [180]. Copper oxide nanoparticles can deliver miR-29b to A549 cells [181], and this may inhibit lung tumor cells' migration, invasion, and metastasis [182]. Taken together, nano-miRNA inhalation might be a new strategy to deal with lung cancer metastasis, with few potential off-target effects and systemic toxicities [183].
Presently, despite the great progress that has been made in traditional lung cancer treatments, there are still multifaceted limitations impairing these treatments' effects, such as therapeutic resistance and drug toxicity. As mentioned above, miRNAs and lncRNAs are widely involved in TGF-β signaling-regulated lung cancer metastasis. Targeting the miRNA/lncRNA-TGF-β signaling axis might provide new insights to decrease lung cancer metastasis and improve the prognosis of lung cancer patients. However, these studies are performed mostly on NSCLC cell lines, and studies on SCLC cell lines and in lung cancer tissues are still lacking. Therefore, further explorations in these microenvironments are required to elucidate the roles of miRNAs and lncRNAs in lung cancer more completely.