Lung cancer is the leading cause of cancer mortality worldwide; however, few diagnostic and predictive biomarkers are available for this devastating disease. Emerging evidence suggests that miRNAs have the potential to regulate translation in a cell cycle-dependent manner, which opens new horizons in advancing our understanding of cancer at the molecular level Citation[1,2].
Single nucleotide polymorphisms & lung cancer susceptibility
In the past 2 years, miRNA studies based on single nucleotide polymorphisms (SNPs) have yielded some copy number variants related to lung cancer. Hu and coworkers found that the SNP (rs11614913) in miR-196a2 is associated with survival in individuals with non-small-cell lung carcinoma (NSCLC) Citation[3]. Survival is significantly decreased in individuals who are homozygous CC at SNP (rs11614913). Binding assays reveal that this SNP can affect binding of mature miR-196a2-3p to its target mRNA; therefore, the SNP (rs11614913) in miR-196a2 may be a prognostic biomarker for NSCLC. In a case–control study, they further found that miR-196a2 rs11614913 variant homozygote CC is associated with a significant increased risk of lung cancer compared with their wild-type homozygote TT and heterozygote TC. These findings suggest that functional SNP (rs11614913) in miR-196a2 can also contribute to lung cancer susceptibility Citation[4]. On the other hand, Chin and coworkers identified the LCS6 variant allele in a KRAS miRNA complementary site significantly associated with an increased risk for NSCLC among moderate smokers Citation[5]. This variant allele represents a new paradigm for let-7 miRNAs in lung cancer susceptibility. Since variations in the sequences of miRNAs can affect how humans develop cancers and respond to therapeutic agents, SNPs are considered to be key enablers in realizing the concept of personalized medicine. Further characterization of miRNA SNPs may open new avenues for the study of cancer and therapeutic interventions.
miRNAs as lung cancer biomarkers
MicroRNAs (noncoding RNA molecules that regulate gene expression) are released by cancer cells and circulate in the biofluids with remarkable stability, which gives them the potential to become a new class of biomarkers to detect cancer at its early stage. Chen and coworkers showed that serum and plasma contain a large amount of stable miRNAs derived from various tissues or organs Citation[6]; the expression profile of these miRNAs shows great promise as a novel noninvasive biomarker for early diagnosis of cancer. Employing Solexa (a high-throughput sequencing technology of small RNAs), they detect two NSCLC-specific serum miRNAs (miR-25 and miR-223). Elevated expression levels of these miRNAs in serum are the blood-based biomarkers of NSCLC, which can be easily detected by quantitative reverse-transcription (qRT) PCR.
Interobserver variability and the lack of specific, standardized assays limit the current abilities to adequately stratify NSCLC patients for targeted therapies. Lebanony and coworkers identified miR-205 as a highly specific marker for squamous cell lung carcinoma Citation[7]. A miRNA-based qRT-PCR assay based on miR-205 expression distinguishes squamous from nonsquamous NSCLC formalin-fixed, paraffin-embedded samples with 96% sensitivity and 90% specificity. This standardized diagnostic assay contributes to the highly accurate subclassification of NSCLC patients.
Prognostic markers help to stratify patients for treatment by identifying patients with different risks of outcome (e.g., recurrence of disease), and are important tools in the management of lung cancer. Using looped qRT-PCR, Markou and coworkers detected overexpression of mature miR-21 and miR-205 in NSCLC tissue specimens Citation[8]. Mature miR-21 overexpression correlates with overall survival of the patients, whereas overexpression of mature miR-205 does not. These results suggest that overexpression of mature miR-21 detected by qRT-PCR is an independent negative prognostic factor for overall survival in NSCLC patients.
Most human cancer deaths are caused by metastasis; identifying the molecular signatures where the expression changed in metastasis offers potential targets to inhibit metastasis. To seek a specific miRNA expression signature characterizing the metastatic phenotype of solid tumors, Baffa and coworkers performed a miRNA microarray analysis on paired primary tumors (including lung cancer) and one of their related metastatic lymph nodes Citation[9]. They identified a metastatic cancer miRNA signature comprising 32 dysregulated miRNAs, and some of them have a well-characterized association with cancer progression, such as miR-10b, miR-21, miR-30a/e, miR-125b, miR-141, miR-200b/c and miR-205. Their results suggest that miRNAs may represent a novel diagnostic tool in the characterization of metastatic cancer gene targets.
Therapeutic potential for miRNAs
Increasing numbers of studies show that miRNA expression correlates with various cancers and some miRNAs function as oncogenes and tumor suppressors; these accumulated understandings of the underlying mechanism provide opportunities towards the design of molecular medicines based on miRNA modulation. Du and coworkers showed that exogenous overexpression of miR-93, miR-98 and miR-197 inhibits Fus1 protein expression, leading to lost or greatly diminished tumor-suppressor function in small-cell lung cancer (SCLC) Citation[10]. Individual deletion of these miRNA target sites in the FUS1 3´ untranslated region restores the expression level of Fus1 protein. Their results suggest that miRNAs can regulate expression of tumor-suppressor gene in lung cancer. Ebi and coworkers also found frequent overexpression of miR-17–92 in SCLC cells Citation[11]. This miR-17–92 overexpression induces γ-H2AX, which reflects continuing DNA damage in Rb-inactivated SCLC and, consequently, leads to genetic instability. Therefore, this miRNA cluster may be a therapeutic target candidate for SCLC.
In the study of NSCLC, Garofalo and coworkers showed that levels of miR-221 and miR-222 are increased in TNF-related apoptosis-inducing ligand (TRAIL)-resistant NSCLC cells Citation[12]. These miRNAs interfere with TRAIL signaling through targeting the 3´ untranslated region of p27(Kip1) mRNA. They demonstrated that miR-221 and miR-222 impair TRAIL-dependent apoptosis by inhibiting the expression of key functional proteins, thus making these miRNAs as diagnostic tools or therapeutic targets for TRAIL-resistant NSCLC.
On the other hand, Weiss and coworkers showed that miR-128b directly regulates EGF receptor (EGFR) in NSCLC cells Citation[13]. This miRNA loss of heterozygosity is frequent in NSCLC patient samples and correlates significantly with clinical response and survival following gefitinib. By contrast, the expression and mutation status of EGFR does not correlate with survival outcome. These results suggest that identifying miRNA regulators of oncogenes can have far-reaching implications for lung cancer patients, including development of early biomarkers, improving patient selection for targeted agents, or development of novel therapeutics. Mutation in EGFR is more frequent in never-smoking patients with NSCLC. Cho and coworkers found ectopic expressions of six miRNAs (miR-21, miR-126*, miR-145, miR-182, miR-183 and miR-210) in the lung cancer tissues from this subtype of patients Citation[14]. They showed that restoration of miR-145 inhibits cancer cell growth in EGFR mutant NSCLC. Their results suggest that miRNA study may identify useful diagnostic markers and potential therapeutic targets for a special subset of lung cancer. Seike and coworkers also identified aberrantly expressed miRNAs in cases of never-smoker lung cancer, including miR-21 and miR-138Citation[15]. The changes in expression of miR-21 are more remarkable in cases with EGFR mutations than in those without these mutations. Antisense inhibition of miR-21 not only enhances EGFR-tyrosine kinase inhibitor-induced apoptosis in never-smoker NSCLC with mutant EGFR, but also induced apoptosis by itself in never-smoker NSCLS with wild-type cases.
Some miRNAs have been identified to demonstrate a tumor-suppressor role in lung cancer. Wu and coworkers found that Ubc9 is upregulated in various cancer specimens, including lung cancer Citation[16]. This E2-conjugating enzyme plays a vital role in sumoylation-mediated cellular pathways, ultimately impacting cell growth and cancer development. Ectopic expression of miR-30e negatively regulates Ubc9 expression in cancer cells and suppresses cell growth. They showed that Ubc9 is a direct target for miR-30e. On the other hand, Liu and coworkers found that miR-34c, miR-142–5p and miR-145 are repressed in transgenic lung cancers Citation[17]. Individual overexpression of these miRNAs significantly represses lung cancer cell growth, whereas anti-miRNA cotransfections antagonized this inhibition. Transfection of miR-34c represses cyclin E and provides a mechanism for observed growth suppression. Bandi and coworkers also showed that miR-15a and miR-16 are frequently deleted or downregulated in NSCLC Citation[18]. Physiologic concentrations of these miRNAs directly regulate cyclins D1, D2 and E1. However, NSCLC cells lacking Rb are resistant to cell cycle arrest induced by them, whereas reintroduction of functional Rb resensitizes these cells to miRNA activity. Their results indicate that miR-15a and miR-16 are implicated in cell cycle control in an Rb-dependent manner and likely contribute to the tumorigenesis of NSCLC. In another study, Kumar and coworkers demonstrated that ectopic expression of let-7g in KRAS(G12D)-expressing lung cancer cells induced both cell cycle arrest and cell death Citation[19]. The expression of let-7g induced from lentiviral vectors substantially reduces lung tumor burden in an autochthonous mouse model of NSCLC in vivo. This let-7g-mediated tumor suppression is more potent in lung cancer cells harboring oncogenic KRAS mutations than in cells with other mutations.
Perspectives & challenges
Rapid advances in platform technologies, such as SNP analysis, genome-wide transcriptional profiling, miRNA microarray, BeadArray, and other ’omics technologies offer the potential for revolutionary development in the study of miRNAs in cancer Citation[20–24]. Although the biological importance of miRNA is becoming increasingly apparent, regulation of miRNA expression in lung cancer is not fully understood. There are some questions that must be addressed. As miRNAs have an important role in self-renewal and differentiation of embryonic and tissue-specific stem cells, do they also have a regulatory role in lung cancer stem cells? Promising findings of a lung cancer-associated miRNA in one study is not adequate to support a solid report; multicenter and an independent cohort of studies would be needed to crossvalidate the discovery. MiRNA expression studies in lung cancer indicate their importance and potential use as disease classifiers and prognostic tools in this field but the true clinical utility and the limits of this new application are yet to be established. Although miRNA-based therapeutics in several proof-of-principle experiments have demonstrated exciting effects, the developing progress of antisense or siRNA drugs has been hampered by delivery and stability problems. Even the newly invented locked nucleic acid technology presents a breakthrough in oligonucleotide carrier; results from a Phase I trial are still pending to confirm the use of this antagomir in respect to safety for human application Citation[25]. Nevertheless, the rapid progress in miRNA studies points to their tremendous potential in the diagnosis and prognosis of lung cancer, as well as their potential role in future therapeutics.
Financial & competing interests disclosure
The author has no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.
No writing assistance was utilized in the production of this manuscript.
Related Research Data
References
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