LncRNAs in Cancer Development

Abstract

The goal of this chapter is to make an overview of the identified changes in lncRNAs expression levels accompanying cancer development. In general, the statistics allows us to establish a fact of association between the phenomenon and the process, but not to discover underlying mechanisms. In the context of the discussed topic, the phenomenon and the process are changes in lncRNA expression levels and cancer development. However, the underlying mechanisms, allowing such changes as in lncRNAs expression levels to have an impact on the cancer development, mostly remain uncertain and not clear. The first part of the chapter aims to shortly highlight the possible mechanisms of lncRNAs` impacts on the main processes of cancer development, like EMT, cancer cell progression, invasion, and metastasis. The second part examines in more detail the role of lncRNAs in some of the main nosology of oncological diseases: The participation of lncRNAs in the formation and development of breast cancer is considered, the main aspects of the importance of lncRNAs in lung cancer are presented, and studies on the participation of lncRNAs in the formation of colorectal cancer are described.

1. Introduction

The protein-coding part of genomes, and the human genome is no exception, is not the least, but quantitatively an insignificant part of it: About 1.9% of transcribed RNA is responsible for protein coding, and in the mammalian genome, only 70% of RNA is transcribed [1]. A significant part of RNA is not protein coding, which raises the question of its role in the human beings and alive organisms in general. These non-coding RNAs are represented by molecules of different sizes and are classified according to the number of nucleotides and not according to their functional properties [2].

For a non-coding RNA of approximately 200 nucleotides in size, recent researches have elucidated many functions and mechanisms. It was experimentally demonstrated that the functions of such RNAs are associated with the mechanisms of DNA regulation, with epigenetic control of gene expression. These RNAs are involved in RNA maturation, and indirectly in gene regulation, for example, through interaction with promoters’ sites. The following functions of non-coding RNAs have been identified: the inactivation of the X9 chromosome, formation and maintenance of nuclear architecture, and its importance in imprinting [3]. Small nuclear RNAs (snRNAs), as studies have shown, are involved in splicing processes, while transport RNA is responsible for the transport and identification of triplet nucleotides. Ribosomal RNAs, being the most represented in cells, are involved in the processes of formation of the ribosomal framework. Small nuclear RNAs (snoRNAs) perform, for example, functions related to housekeeping RNA chemical transformation processes [3]. In addition, there are many types of small non-coding RNAs involved in gene regulation, like miRNAs, which are 20–25 nucleotides and directly or indirectly regulate gene expression. Among these molecules are piRNAs (piwi-interacting RNAs), which differ from miRNAs in biogenesis and are mainly responsible for the targeting and switching off the expression of transposable and repetitive elements to support and maintain the constancy and stability of the embryonic genome [4]. The last most studied are siRNAs, which are involved in such processes as chromatin organization, control of mobile genetic elements, and gene regulation at the translation and at the post-translational level, and are also associated with the function of maintaining genome stability in some cells [4, 5].

Long non-coding RNAs (here and further lncRNAs) often contain a poly-A tail and can also be spliced, just like mRNA. It is worth noting that for a large number of lncRNAs, the functional significance has not yet been determined (and the exact genomic annotations have also not been identified). It is estimated that such lncRNAs constitute between 5400 and 10,000 of the lncRNA transcript pool. But still, the identification of mutations in the genes responsible for encoding those lncRNAs that are involved in the formation of cancer has made it possible to significantly rethink the molecular genetics of cancer development [6, 7].

In general, long and small non-coding RNAs are involved in the development of various diseases, including the development of cancer. lncRNAs are involved in processes such as angiogenesis, differentiation, and cell proliferation. They are also involved in apoptosis and cell migration, and are involved in the process of metastasis. lncRNAs can act both as oncogenes and as tumor suppressors [6, 7].

2. Long non-coding RNAs in cancer

Obviously, cancer is a complicated process involving many mechanisms and metamorphizing cell cycle in different steps. LncRNAs, which are expressed in various cell states, regulate gene expression processes so they may reflect the development of cancer. One possible mechanism is the binding of lncRNAs to proteins and the formation of scaffolds that are involved in the construction of regulatory complexes. One example of the functioning of such complexes is HOTAIR; it is involved in the trimethylation of LSD1, as well as H3K27, due to which demethylation of H3K4me2 occurs. HOTAIR implements this function through interaction with the PRC2 complex [8]. Qian et al. admit two other possible mechanisms of gene expression regulation: The first is the functioning of lncRNA as a decoy, which prevents the binding of transcription regulators. According to this theory, lncRNAs may bind miRNAs and prevent RNA degradation competing endogenous RNAs. The second possible way is guiding the transcription factors to specific sites like MEG3, which guides PRC2 and forms a complex with DNA [9].

Epithelial-mesenchymal transition or EMT is one of the key processes of cancer formation in epithelial cells and besides all cell types, while the main event of EMT is the total imbalance of epithelial cells following loss of the corresponding architecture, so anterior polarity and posterior polarity arise, apical-basal polarity is lost, and the shape of cells changes, what partly happen due to the damage to the structure of the cytoskeleton. These hallmark events are accompanied with disruption of epithelial gene expression markers that allow the determination of mesenchymal phenotypes along with cell growth and movement function changes. The new phenotype they acquire is often characterized with the possibility of extracellular matrix (ECM) protein degradation and apoptosis and aging ignoring [10]. All these events are realized through a majority of different molecular mechanisms, as it was already mentioned above, including disruption of signal pathways and changes in gene expression and regulation, and lncRNAs in this sense may play a pivotal role in the processes underlying EMT as well as in promotion cell invasion and migration. When examining lncRNAs in detail, it can be seen that a set of molecular processes occurring around them can significantly influence the functions of lncRNAs. In addition, depending on these processes, the functions of lncRNAs can become completely opposite. Liu et al. highlight the inconsistency of functions of H19, one of the earliest described lncRNAs63. In bladder cancer cells, it associates with the polycomb repressive complex 2 (PRC2) component EZH2, leading to decreased E-cadherin by increasing the activity of β-catenin, supporting the migration of malignant cells in vitro and a transition toward a mesenchymal state. Simultaneously, Liu mentioned studies in which H19 interacted with miR-200c, as well as h miR-200b. As a result of this interaction, GIT2 is suppressed, which in turn stimulates the formation of metastatic colonies. At the same time, in circulating tumor cells, H19 interacts with microRNA let-7b and blocks it, resulting in the suppression of CYTH3. Suppression of CYTH3 promotes the formation of a mesenchymal phenotype [11]. Along with this, lncRNAs are involved in the promotion of invasion and migration of tumor cells, so NORAD decrease has a significant correlation with lymph node metastases from patients with lung and breast cancer, while ectopic expression of this lncRNA in vitro inhibited invasion and migration [12]. The main mechanisms of action of this lncRNA on carcinogenesis and the development of metastases include suppression of S100P functions, due to which S100P-mediated signaling, which activates invasion and formation of metastases, is disrupted [12].

LncRNA CCAT2, through interaction with TCF7L2, activates MYC transcription and WNT signaling. Increased expression of TCF7L2 occurs in microsatellite-stable and metastatic CRC [13]. Ling showed that miR-20a and miR-17-5p are also activated by CCAT2 through TCF7L2-mediated transcriptional regulation, with CCAT2 being a downstream target of WNT, which may support a feedback mechanism. In addition, CCAT2 expression has been shown to be influenced by SNP status: The G risk allele produces more CCAT2 transcripts [13].

Jessica Silver-Fisher et al. found 148 differentially expressed RNAs associated with metastasis (RAMS). The results were obtained by sequencing the transcriptome of normal tissues, primary cancer tissues, and distant metastases. The main focus was on RAMS11 as it was most associated with poor disease-free survival. Additionally, RAMS11 has been shown to promote aggressive phenotypes in both in vitro and in vivo studies. The experiments presented in the work revealed the mechanism of RAMS11-dependent recruitment of Chromobox protein 4 (CBX4)—this protein is an activator of topoisomerase 2 alpha (TOP2α). RAMS11 expression was found to be highly overexpressed in liver metastases from colorectal cancer patients compared with its expression in primary tumor cells [14].

LncRNAs may participate in the activation of proliferation and processes underlying insensitivity to growth factors. LncRNAs may be involved in different mechanisms of proliferation activation such as activating the signal receptors or like PVT1 being involved in the regulation of receptor abundance. Some lncRNAs are involved in providing resistance to expression suppressors and tumor growth inhibitors by affecting different stages of translation and transcription. LincRNA-p21 and p21 (inhibitor of CDK2) are involved in the initiation of transcription by the scaffolding of transcription factor complexes, and gadd7 and Cdk6 participate in the destabilization of mRNA transcripts and thus modify transcription elongation [15].

Due to new technologies development, it became evident many lncRNAs are connected to carcinogenesis, making them new perspective players into the field of molecular biology. The main function of lncRNA is to regulate the expression of other genes, which gives these molecules a critical role in the existence of cells. Undoubtedly, the discovery of new biomarkers that allow the assessment of various aspects of oncological diseases and open up new opportunities for diagnosis, prognosis, and treatment is the most important task of modern medicine. In this regard, the study of molecules such as lncRNAs can be a promising approach. Next, we will focus and consider the involvement of lncRNAs in the development of breast cancer, lung cancer, and colorectal cancer.

3. LncRNA in breast cancer

Currently, breast cancer is the second leading cause of death among women worldwide, while being the most common cancer [16]. Recent studies have shown that lncRNA expression changes are associated with disease progression and survival in breast cancer patients [17, 18, 19, 20]. Xu et al. showed that lncRNA 179 and lncRNA 216 have an association with the overall survival of breast cancer patients (P values <0.05 for both). In their study, they demonstrated significant changes in the expression levels of 10 lncRNAs associated with overall survival. The work included the following lncRNAs: MAPT-AS1, RP11-120 K18.2, SPACA6P-A, AC025016.1, LINC02037, SPACA6P-AS, RP11-616 M22.1, 7, RP1-37C10.3, RP11-454P21.1, and P11-344E13.4. To identify the relationship with the survival of breast cancer patients, the researchers conducted a univariate Cox regression analysis, which showed for 6 lncRNAs a higher regression coefficient and a decrease in overall survival. A protective role was revealed for lncRNA MAPT-AS1—the expression level of this lncRNA correlated with survival rate in breast cancer patients [17]. In 2021, Qiao et al. demonstrated the effect of lncRNA408 on breast cancer progression, which was significantly enhanced in breast cancer cells undergoing EMT and in breast cancer tumor cells with lymph node metastasis compared to samples without lymph node metastasis [18]. Since lncRNA-408 acts as a sponge for miR-654-5p, it may be partially involved in mitigating miR-654-5p repression of its target LIMK1. This molecular mechanism is partially responsible for the development of breast cancer cells and their metabolic changes, since LIMK1, a gene involved in cytoskeleton stabilization, is one of the target genes of miR-654-5p. In addition, due to a decrease in the expression level of miR-654-5p, the expression of COL1A1, as well as MMP2 and ITGB1, increases [18].

Another lncRNA involved in the development of cancer, especially breast cancer, is the ST7-AS1 lncRNA. In addition to stimulating breast cancer, it may influence the development and progression of laryngeal squamous cell carcinoma. This is due to the involvement of the following molecular mechanisms that control the IL6/JAK STAT3 signaling pathway. This signaling pathway is generally associated with poor prognosis and is frequently activated in many cancers [19]. Reduced expression of lncRNA ST7-AS1 in breast cancer correlates with clinical and advanced pathological characteristics (high grade, HER2 status, age, advanced stage, menopausal stage, histology), poor prognosis, and survival time. GO analysis showed that lncRNA ST7-AS1 is associated with T helper cells and DC cells [20].

Yi et al. found the influence of SLC26A4-AS1 on breast cancer development: In breast cancer patients, decreased SLC26A4-AS1 expression correlated with tarragon receptor status, age, and menopausal status. Low expression was also associated with poor overall survival. SLC26A4-AS1 has been shown to be involved in multiple biological processes required for cancer development, such as protein export, cristae formation, the endosomal sorting complex required for histone modification and transport, ATP generation, and others [21].

DRAIC, also known as LOC145837 and RP11-279F6.1, along with breast cancer has been shown to affect both lung cancer and prostate cancer. Briefly, the expression of this lncRNA in tumor cells was identified as estrogen-dependent and elevated relative to that of neighboring non-cancer cells. Zhao et al., working with RNA sequencing data from the Tumor Genome Atlas, analyzed more than 800 breast cancer samples compared with about 100 normal cell samples and found that the expression level of lncRNA DRAIC was associated with progesterone and estrogen receptors, and also with human epidermal cells [22].

Fonseca-Montaño et al. worked with the BRCA-TCGA and GEO-GSE96058 datasets and found that in patients with luminal B breast cancer, LINC00426 expression levels correlated with overall survival. Further studies showed that LINC00426 expression is associated with various oncological processes and immune pathways in breast cancer. In addition, the expression of LINC00426 was associated with the expression of genes with cytolytic activity, the expression of immune checkpoints, and the level of infiltration of various populations of immune cells [23].

Collina et al., working with biological samples rather than ready-made datasets, found that lncRNA HOTAIR, one of the first to be linked to cancer, was implicated in breast cancer development. Experiments have demonstrated that the expression level of lncRNA HOTAIR is associated with metastases to lymph nodes. In addition, the study revealed that the expression levels of HOTAIR lncRNA have a direct correlation with the expression levels of androgen receptors in samples obtained from patients with triple-negative breast cancer. Based on the data obtained in this way, one can assume a prognostic role of HOTAIR in TNBC, as well as its possible indirect participation in the regulation of the androgen receptor pathway [24].

Gene expression is regulated by lncRNA through different molecular mechanisms. Among them are chromatin modification, translation, and transcription. All these make LncRNA an important biomarker of the oncological process, reflecting those properties of tumor cells that allow them to metastasize, form drug resistance, acquire the ability to invade, vascularize, and progress.

A year before, Li et al. demonstrated, that another lncRNA, lncATB, was shown to be involved in breast cancer development and to be increased in breast cancer cells. The results of experiments on lncATB made it possible to assume its significant contribution to oncological processes, such as invasion, cell migration, and the ability to form clonal cells. Data were obtained from both in vitro and in vivo studies. lncATB has the ability to bind microRNAs of the miR-200 family, due to which it increases the expression of Twist 1. This, in turn, promotes epithelial-mesenchymal transition [25]. All these have shown that the lncATB expression has a direct positive correlation with tumor size and an inverse correlation with the duration of relapse-free survival and overall survival [25].

These lncRNAs, involved in breast cancer development, are listed below in Table 1 and are shown in Figure 1.

#	LNC	Materials	Methods	Prognostic/diagnostic value	Reference
1	RP11-616 M22.1, LINC02037, RP11-344E13.4, RP11-454P21.1, SPACA6P-AS, MAPT-AS, RP1-37C10.3	Biomaterial under study: breast cancer cells—913, non-tumor cells adjacent to cancer—cancer cells—113. Data: clinical data, training data (n = 608), test data set (n = 305).	Illumina HiSeq RNA-Seq	For a panel of 7 lncRNA markers, the ROC curve area was 0.771 for predicting 3-year survival. For a panel of 7 lncRNA markers, the ROC curve area was 0.780 for predicting 5-year survival.	[17]
2	LNC408	BC tissues in patients with BC and lymph node metastasis (N = 35) and with BC and without lymph node metastasis (N = 25) (stage I (N = 19), stage II (N = 18), stage III (N = 23)).	qRT-PCR	Lnc-408 was significantly increased in BC tissues with lymph nodes compared with BC tissues without lymph nodes (P < 0.0001). Lnc-408 expression level was directly related to N stage, TNM stage, and histological grade.	[18]
3	LncRNA ST7-AS1	15 PCR samples; clinical data on breast cancer from the TCGA project; 1065 samples HTSeq-Counts and HTSeq-FPKM.	RT-PCR. TCGA data	Compared with 1065 BC tissues, lncRNA ST7-AS1 expression was significantly higher in 111 healthy cells (p < 0.001). The low expression of lncRNA ST7-AS1 in breast cancer was correlated with a number of clinical characteristics: age, menopause, HER2 status, stage, histological type, survival, and poor prognosis.	[20]
4	SLC26A4-AS1	1109 tissues from patients with breast cancer; 113 normal breast tissue; clinical and RNAseq data from the TCGA project	Single-gene differential analysis of SLC26A4-AS1 and GSEA	The expression level of SLC26A4-AS1 in breast cancer tissues is lower than in normal breast tissues (0.267 ± 0.015 vs. 0.808 ± 0.062, P < 0.001, AUC was 0.805). SLC26A4-AS1 expression correlated with menopausal status, ER status, PR, and age.	[21]
5	DRAIC (also known as LOC145837 and RP11-279F6.1)	828 tumor tissues and 105 normal breast tissues from TCGA		In patients with advanced stages of cancer, the expression of lncRNA DRAIC is higher than in patients with early stages. In patients with lymph node metastases, the expression of lncRNA DRAIC is higher than in patients without lymph node metastases. Higher DRAIC predicts shorter overall survival of patients.	[22]
6	LINC00426	BRCA-TCGA (TCGA database through cBioPortal) and GEO-GSE96058 (GDC Data Portal datasets). A total of 927 patients with breast cancer: 490 patients with luminal type A breast cancer; 192 patients with breast cancer luminal type B; 77 patients with HER2-positive breast cancer; 168 patients with bilateral breast cancer. A total of 3052 patients—validation group from the GEO database: 1657 patients with luminal type A breast cancer; 729 patients with breast cancer luminal type B; 327 patients with HER2-positive breast cancer; 339 patients with bilateral breast cancer.	Comparison of gene expression between groups of breast cancer patients with high and low expression of LINC00426 was performed using the R package DESeq2 (version 1.38.1). Values less than 10 were filtered out. RNA-seq data from breast cancer patient cohorts with high and low LINC00426 expression were assessed using GSEA (version 4.1.0).	Patients with non-luminal breast cancer showed increased expression of LINC00426 compared to patients with luminal breast cancer. It was also noted that LINC00426 expression has a prognostic value for overall survival in patients with luminal subtype B breast cancer with PAM50 from the TCGA cohort.	[23]
7	LncRNA HOTAIR	A total of 163 patients who underwent a mastectomy, quadrantectomy, or metastectomy.	HOTAIR in situ was determined using RNAscope.	HOTAIR lncRNA expression is associated with lymph node metastasis and ductal histological type. No association was found between lncRNA HOTAIR and tumor size, proliferation, distant metastases, and overall and disease-free survival.	[24]
8	LncRNA ATB	A total of 131 samples from patients with primary breast cancer following surgery. Sixteen samples of normal breast tissue. None of the patients received radiation therapy, endocrine therapy, chemotherapy, or targeted therapy.	Expression analysis of lncRNA EMT and lncATB was performed using real-time PCR (the study was performed in triplicate using specific primers). Normalization was performed using the 2-ΔΔCt method; the expression level of β-actin was used for normalization. To investigate the molecular mechanism, biotin-labeled lncATB probes were used and incubated with cytoplasmic lysate of the MCF-7 cell line. The resulting lncRNA-miRNA complexes were then collected using specific magnetic beads. qRT-PCR was used to detect miR-200.	In cells of triple-negative and luminal breast cancer B (Her-2-positive), there are increased levels of expression of lncRNAs LncATB and Twist1. Luminal type A breast cancer cells have lower expression levels of lncRNAs LncATB and Twist1. In normal breast cells, the expression levels of lncRNAs LncATB and Twist1 are the lowest. For lncATB, the AUC was 0.851. For Twist1, the AUC was 0.778. OS and DFS are longer in the low lncATB expression group than in the high lncATB expression group. The binding of lncATB to the miR-200 family has been demonstrated.	[25]

Table 1.

LncRNA and breast cancer.

Figure 1.

LncRNAs in breast cancer development.

4. LncRNA in lung cancer

LncRNAs, as in other cancers, play a significant role in lung cancer. LncRNAs PVT1 and also MALAT1 can be noted—their expression is often increased in lung tumors. Blocking these lncRNAs, or vice versa, activation, affects proliferation and cell growth [26]. LINC00680 is another lncRNA implicated in the development of lung cancer. This lncRNA binds to GATA6, LINC00511, and the chromatin-converting enzyme EZH2, thereby suppressing the tumor suppressor genes p57 and LATS2, resulting in the development of non-small-cell lung cancer [26].

Hu et al. in 2016 demonstrated a promising potential of lncRNAs as non-invasive markers in lung cancer: They showed a high diagnostic value of these molecules working with plasma lncRNAs. From 21 previously identified lncRNAs, they identified SPRY4-IT1, ANRIL, and NEAT1 as the targets to be validated by qRT-PCR. The lncRNAs SPRY4-IT1, ANRIL, and NEAT1 were found to be significantly increased in plasma samples from patients with non-small-cell lung cancer. At the same time, ANRIL in the blood plasma of such patients proved to be the most informative diagnostic marker (ROC = 0.798). At the same time, when combined with lncRNA SPRY4-IT1 and NEAT1, the diagnostic significance increases (specificity 92.3%, sensitivity 82.8%) [27]. The training set included an independent group of 20 plasma samples from patients, and the validation set included 50 NSCLC patients and 50 different independent groups, allowing study control to confirm apparent differences in relative expression levels of biomarker candidates and 50 healthy controls, respectively [27].

Esfandi et al. obtained the data for NEAT1 expression in lung cancer with same tendencies as in the study by Hu et al. mentioned above. However, they worked with lncRNAs isolated from tissue samples, not from plasma, and showed that lncRNAs such as PVT1, HOXA transcript antisense RNA myeloid-specific 1 (HOTAIRM1), Fas-antisense 1 (FAS-AS1), taurine-regulated gene 1 (TUG1), Growth arrest-specific 5 (GAS5), copy nuclear paraspeckle accumulation 1 (NEAT1), TNFα, and hnRNPL-related immunoregulatory LincRNA (THRIL) have diagnostic and prognostic potential. Changes in their expression levels were shown by Esfandi et al. in 32 NSCLC samples and their corresponding non-cancerous cells [28]. It is interesting to admit a gender-dependent expression in some lncRNAs: NEAT1 showed a significant increase in NSCLC cells in men compared to the corresponding non-cancerous cells, and this change was not repeated in women and these data were confirmed in other published data of Pan et al. in 2015 and Hu et al. in 2016. Conversely, in non-small-cell lung cancer cells from male and female patients, FAS-AS1 expression was significantly reduced compared to corresponding normal lung cells. The expression levels of HOTAIRM1, as well as the expression levels of GAS5, TUG1, and THRIL, are significantly reduced in non-small-cell lung cancer cells compared with normal lung cells from male patients.

Increased expression of lncRNA RMRP is also characteristic of lung cancer. Meng et al. analyzed lncRNA expression in lung adenocarcinoma samples as well as in adjacent normal tissue samples (25 of 35). Experiments showed that in 71% of cases in adenocarcinoma samples, the expression level of lncRNA RMRP was increased compared to normal tissues. RMRP expression was also found to be increased in lung adenocarcinoma cell lines compared to bronchial epithelial cell lines. A possible molecular mechanism of RMRP may be inhibition of miR-206. Experiments with the H1299 cell line showed that cell proliferation and migration induced by RMRP are suppressed when miR-206 expression is restored [29].

Ren et al. [20] showed not only the association between the expression level of lncRNA and cancer development but also the association of mutations in lncRNAs sequences and cancer development. HOTAIR, which is known to be increased in a variety of cancers, was investigated in association with three SNPs identified with the MassArray system and these SNPs were rs920778, rs1899663, and rs4759314. In these studies, Ren et al. found that these SNPs were not significantly correlated with either lung cancer type, disease severity, metastases to lymph nodes, or cancer stage. There were also no correlations between these SNPs and family history. There is little gender dependence: using gender stratification, and the AG genotype compared to the AA genotype for rs920778 and the AG + GG genotypes for rs920778 were shown to be protective factors against NSCLC in women. In the analysis of smoking, it was shown that the AG + GG genotype was a protective factor against NSCLC in non-smokers compared with the AA genotype rs920778. In case with rs1899663 and rs4759314 genotypes, no statistical differences were demonstrated in both gender and smoking stratifications [30].

Lin et al. in 2015 obtained the same data about ANRIL overexpression in lung cancer as in the aforesaid study by Hu et al. Lin et al. analyzed ANRIL expression in 87 NSCLC tissues and three lung cancer cell lines. In tissue samples from patients with non-small-cell lung cancer, the expression level of lncRNA ANRIL was increased compared to samples from adjacent non-tumor tissues. In addition, the expression level of lncRNA ANRIL was correlated with higher-stage and late lymph node metastasis. It is noted that lower overall survival is observed in groups with high expression of lncRNA ANRIL. In addition, when the expression of lncRNA ANRIL is blocked, the proliferation, migration, and invasion of cancer cells in vitro are observed [31].

These lncRNAs, involved in breast cancer development, are listed below in Table 2 and in Figure 2.

#	LNC	Materials	Methods	Prognostic/diagnostic value	Reference
1	lncRNAs such as SPRY4-IT1, CCAT2, RGMB-AS1, ANRIL, NEAT, PCAT1.	Plasma samples NSCLC (n = 120), plasma healthy controls (n = 120).	qRT-PCR	SPRY4-IT1, ANRIL, NEAT were significantly increased in NSCLC compared to samples obtained from normal lung cells.	[27]
2	FAS-AS1, GAS5, HOTAIRM1, NEAT1, TUG1, and TNFα - associated with hnRNPL	32 tissue samples from patients with non-small-cell lung cancer. No patient received either radiotherapy or chemotherapy.	Quantitative PCR. ANCT expression level was used for normalization. Expression was assessed by Efficiency^∆CT.	Expression levels of HOTAIRM1, GAS5, TUG1, and THRIL, normalized by ANCT expression, are increased in tissue samples from men with NSCLC compared with normal tissues. The expression level of lncRNA FAS-AS1, normalized by ANCT expression, is significantly reduced in tissue samples from women with NSCLC compared with normal tissues. The expression level of lncRNA FAS-AS1, normalized by ANCT expression, is significantly reduced in tissue samples from men with NSCLC compared with normal tissues. The expression level of lncRNA NEAT1, normalized by ANCT expression, does not change in tissue samples from women with NSCLC compared to normal tissues. The expression level of lncRNA NEAT1, normalized by ANCT expression, is significantly increased in tissue samples from men with NSCLC compared with normal tissues.	[28]
3	LncRNA RMRP	35 lung cancer tissue samples. 35 normal tissue samples (taken next to lung cancer tissue).	Real-time PCR. mRNA expression was normalized to GAPDH and lncRNA expression was normalized to 18S rRNA.	In 25 of 35 lung adenocarcinoma tissue samples (71%), RMRP expression was increased. Ectopic expression of RMRP increased lung adenocarcinoma cell proliferation, colony formation, and invasion.	[29]
4	LncRNA HOTAIR	Blood samples from patients with lung cancer—196 samples. Blood samples from healthy individuals who visited the hospital during the same period but did not have cancer or lung disease—196 samples.	GEPIA (Gene Expression Profile Interactive Analysis) resource was used to evaluate expression. MassArray system was used for analysis of HOTAIR lncRNA SNPs (rs920778, rs4759314, and rs1899663). Multiplex PCR—used to analyze 10–20 ng of genomic DNA. The 384-element SpectroCHIP array was used for locus-specific single-base extension reactions.	HOTAIR expression did not show a significant correlation with overall survival of patients with lung adenocarcinoma. There was also no significant correlation between the level of HOTAIR expression and the prognosis of the disease in male patients with lung adenocarcinoma and squamous cell lung cancer, as well as in female patients with lung adenocarcinoma and squamous cell lung cancer.	[30]
5	lncRNA ANRIL	Tissue samples from patients with non-small-cell lung cancer—87 samples. Lung cancer cell lines are SPC-A1, A549, and NCI-H1650. The cell line of normal human bronchial epithelial cells is 16HBE.	qRT-PCR	In tissue samples from patients with non-small-cell lung cancer, the expression level of lncRNA ANRIL was increased compared to samples from adjacent non-tumor tissues. In addition, the expression level of lncRNA ANRIL was correlated with higher-stage and late lymph node metastasis.	[31]

Table 2.

LncRNA and lung cancer.

Figure 2.

LncRNAs in lung cancer development.

5. LncRNA in colorectal cancer

Ling et al. [13] and Nissan et al. [32] conducted a series of experiments and were among the first to obtain data demonstrating the general significance and possible effects of lncRNAs in colorectal cancer. LncRNA CCAT1, as well as lncRNA CCAT2, is located on chromosome 8q24 [32, 33]. Wu et al. in 2023 published a large review on the role of lncRNAs in colorectal cancer. This review cited data from many studies that showed that the expression levels of lncRNAs CCAT1 and CCAT2 are significantly increased in samples obtained from tumor tissues compared with the expression level in normal tissues. In addition, high expression levels of these lncRNAs were correlated with poor prognosis. In addition to these lncRNAs, HOTAIR, H19, and MALAT-1 are also significant for colorectal cancer [34].

Ye et al. showed that NALT1, which has been implicated in the occurrence of gastric cancer (GC), was overexpressed in 76 CRC patients ranging from stages I to IV. Increased NALT1 expression is associated with advanced-stage cancer. Inhibition of NALT1 reduced cell proliferation, invasion, and migration. On the contrary, when NALT1 expression increased, cell proliferation, invasion, and migration were activated both in vitro and in vivo. Studies have shown that the putative molecular mechanism of action of NALT1 is realized through the binding of miRNA-574-5p, due to which the expression of PEG10 is increased [35].

Another lncRNA with a role in colorectal cancer is BCYRN1. In addition, studies have shown that high BCYRN1 expression promotes metastasis in colorectal cancer patients and is also generally associated with poor prognosis. In an in vitro study, experiments also showed that downregulation of lncRNA BCYRN1 using si-BCYRN1 resulted in a significant reduction in invasion and migration [36].

Just like lncRNA-408 for miR-654-5p or lncRNA-ATB for miR-200 or H19 for miRNA let-7b, lncRNA cancer susceptibility 15 (CASC15) binds to miRNA-4310 acting like a sponge. lncRNA CASC15, due to the suppression of microRNA-4310, increases the expression of LGR5, due to which the Wnt signaling pathway is activated. Due to the activation of the Wnt signaling pathway, there is an increase in cell proliferation, invasion, and migration of colon cancer cells. It has also been demonstrated that CASC15 expression is increased in colorectal cancer cells and is associated with metastasis and clinical stage [37]. These are examples of the formation of the lncRNA-microRNA-mRNA signaling axis in which lncRNA 15 and SURC play an important role. The lncRNA-miRNA mechanism in colorectal cancer was demonstrated by the study by Yu et al., where novel lncRNA-miRNA mechanism in colorectal cancer was described and where it was shown that CCAT2 can selectively inhibit miR-145 growth by inhibiting pre-miR-145 by Dicer and its export to the cytoplasm [38].

As it was mentioned above, another lncRNA-miRNA axis in colorectal cancer, promoting tumor development, is formed by lncRNA SURC, which is overexpressed in this type of cancer. SURC promotes CCND2 expression by inhibiting the expression of miR–185-5p in colorectal cancer cells, thus being a specifically upregulated lncRNA in colorectal cancer cells. Along with this, Li et al. have recently demonstrated that the SURC/miR–185-5p/CCND2 axis may be targetable for colorectal cancer diagnosis and therapy [39]. Changes in SURC expression level were identified while comparing lncRNA expression profiles in cancer and normal tissues in mice models. The elevation of lncRNA-mAK028845 expression and its upregulation during colorectal cancer initiation were shown in AOM/DSS-induced CAC model. However, the use of this lncRNA as cancer-specific is problematic, since, as was later revealed, it is actively expressed in non-cancerous tissues of mice. Sequencing revealed that lncRNA AK028845 (named SURC) is located on chromosome 17 in humans, and it contains an exon of the KRT37 gene and a pseudogene KRT41 [39].

These lncRNAs, involved in breast cancer development, are listed below in Table 3 and in Figure 3.

#	LNC	Materials	Methods	Prognostic/diagnostic value	Reference
1	LncRNA NALT1	76 patients with colorectal cancer from stages I to IV. None of these patients received either radiation therapy or chemotherapy.	RT-qPCR—expression level was used for normalization GAPDH. The studies were carried out in triplets. Expression measured by the 2-ΔΔCt method.	High NALT1 expression correlated with advanced cancer stage. Suppression of NALT1 expression reduced cell proliferation, migration, and invasion. Increased expression of NALT1 resulted in increased proliferation, activation of cell migration, and invasion both in vivo and in vitro.	[35]
2	ncRNA BCYRN1	150 samples of tumor tissue from patients with colorectal cancer after surgery. 150 normal tissues (taken near tumor tissue) from patients with colorectal cancer after surgery.	qRT-PCR	In tissue samples from patients with colorectal cancer, the expression level of lncRNA BCYRN1 is significantly increased compared to adjacent normal tissues, which is also associated with a poor prognosis.	[36]
3	LncRNA CASC15	Samples of tumor tissue and adjacent normal tissue from patients with colorectal cancer after surgical treatment—93.	RT-qPCR, normalization was performed according to the level of GAPDH expression.	An increased level of CASC15 expression led to the activation of the Wnt signaling pathway through mira4310, resulting in an increase in the proliferation of colon cancer cells and increased metastasis.	[37]
4	LncRNA SURC	Samples of tumor tissue from patients with colorectal cancer who underwent surgery—150.	qPCR and FISH	A higher number of SURC-positive cells in colorectal cancer tissues compared with adjacent normal tissues were detected by FISH staining. SURC was significantly more expressed in stage II–III than in stage I patients.	[39]

Table 3.

LncRNA and colorectal cancer.

Figure 3.

LncRNAs in colorectal cancer development.

6. Conclusions

However, the extent and nature of lncRNAs promoting the interlocking pathological hallmarks in breast cancer, lung cancer, colorectal cancer, or any other type of cancer remain unclear. Despite the advantage of lncRNA as a diagnostic marker compared to protein-coding RNAs, which is in their own expression as it is a better indicator of the tumor status, there remain some limitations of the study of lncRNA expression in different cancers. These limitations include its retrospective nature and bioinformatics approach based on transcriptomic data, which requires validation of these findings through methodologies like multiplex immunofluorescence or flow cytometry. This validation process is not always possible to be performed because of the unavailability of biological samples, for example. Anyway, lncRNAs remain a promising approach for biomarker studies with further use in clinical routine practice in cancer diagnosis, monitoring, and treatment.

Conflict of interest

The authors declare no conflict of interest.