The major categories of ncRNAs.
The discovery of short regulatory RNAs has recently directed the attention of scientists to parts of the genome that previously had been regarded as “junk” DNA because they did not encode protein products. The revelation that even protein-noncoding sequences had biological functions began the era of discovering the world of noncoding RNAs (ncRNAs). Of these ncRNAs, microRNAs (miRNAs) and long noncoding RNAs (lncRNAs) are the most numerous and best-known ncRNA groups. miRNAs and lncRNAs are important regulators of hematopoiesis, and their abnormal function has serious implications for phenotypes. Deregulation of these ncRNAs is found in hematopoietic disorders, and they also contribute to the development and progression of myelodysplastic syndromes (MDS). Properties of ncRNAs such as stability and tissue specificity make these molecules highly promising diagnostic and prognostic markers as well as interesting therapeutic targets. This chapter summarizes our knowledge on the contribution of ncRNAs to the pathogenesis of MDS and discusses their potential applicability in disease diagnostics and prognosis.
- myelodysplastic syndromes
- noncoding RNAs
- long noncoding RNAs
Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell (HSC) disorders, characterized by dysplastic and ineffective blood cell production, with a tendency for transformation to acute myeloid leukemia (AML). The diagnosis is generally suspected based on the presence of an abnormal complete blood count and is confirmed by performing bone marrow (BM) aspiration and biopsy, demonstrating morphological evidence of dysplasia. A number of additional tests, including cytogenetics, flow cytometry, and molecular genetics, are needed to complete the laboratory evaluation of patients with MDS .
Because of the large heterogeneity of MDS, the development of additional molecular tools able to refine the prognostic scoring system, to predict outcome, and to monitor the response to treatment is required. Recently, application of new high-throughput methods such as next-generation sequencing (NGS) has identified recurrent somatic mutations in MDS cells. In particular, point mutations in the TP53, EZH2, ETV6, RUNX1, and ASXL1 genes have been shown to be associated with specific clinical features and poor overall survival, independent of established risk factors . Even though approximately 78% of MDS patients carry at least one oncogenic mutation , there is a long list of mutations in more than 50 genes with often unclear etiology, complicating the use of somatic mutations as simple and universal markers of MDS prognosis.
Concerning MDS pathogenesis, substantial progress has been made in recent years. A vast literature has become available regarding the spectrum of cytogenetic abnormalities, gene mutations, epigenetic modifications, gene expression patterns, and deregulated signaling pathways (e.g., apoptosis, proliferation, immune response, chromatin remodeling, RNA-splicing machinery, oxidative damage/DNA repair, microenvironment interactions, and others) associated with the disease. In this review, we discuss the contributions of noncoding RNAs (ncRNAs) to the pathogenesis of MDS as well as their potential applications as novel molecular markers for clinical purposes.
2. Noncoding RNAs
At the end of the last millennium, the importance of noncoding RNAs was completely unknown. Up to that point, the scientific community focused on genes that coded for proteins. The classic dogma of molecular biology postulated that DNA was transcribed into RNA, which was then translated into protein, ignoring all non-protein-coding sequences. Only in 1993 did the importance of miRNAs begin to be revealed. The discovery of the first miRNA, lin-4, from
2.1. Types of noncoding RNAs
Functional ncRNAs can be divided into two main types: infrastructural and regulatory ncRNAs. Infrastructural ncRNAs appear to have housekeeping roles in translation and splicing; they include species such as ribosomal RNAs (rRNAs), transfer RNAs (tRNAs), and small nuclear RNAs (snRNAs) that are involved in splicing events. Regulatory ncRNAs, including long noncoding RNAs (lncRNAs), microRNAs (miRNAs), and piwi-interacting RNAs (piRNAs) are involved in the modification and interactions with other RNAs. However, ncRNAs can also be categorized on the basis of length (small, 18–31 nt; medium, 31–200 nt; and long, >200 nt), structure (circular RNAs (circRNAs)), or subcellular localization (small nuclear RNAs (snRNAs) and small nucleolar RNAs (snoRNAs)). Other infrequent ncRNA types such as trans-spliced transcripts and macroRNAs that comprise enormous genomic distances, or multigene transcripts containing several genes or even the whole chromosome, further complicate efforts at systematic classification . Moreover, clear categorization of ncRNA species is difficult, as many ncRNA transcripts often share properties of several categories. The major classes of ncRNAs are summarized in Table 1.
|rRNA||Ribosomal RNA||RNA that is directly incorporated into the ribosome||Large subunit (5S–121 nt, 5.8S–156 nt, 28S–5070 nt)
Small subunit (18S–1869 nt)
|tRNA||Transfer RNA||Transfer amino acids to the ribosome for protein construction||76–90 nt|
|snRNA||Small nuclear RNA||Small RNA located in the nucleus, involved in spliceosomes (e.g., U1, U2, U5, U4, and U6), RNA modification, and other functions. Also commonly referred to as U-RNAs||∼150 nt|
|snoRNA||Small nucleolar RNA||RNA located in the nucleolus, mostly involved in modification of other RNAs, such as rRNA (C/D-box and H/ACA box snoRNAs) or spliceosomal RNA (scaRNA)||∼60–250 nt|
|miRNA||microRNA||A short single-stranded RNA that usually suppresses the translation of target mRNA by binding to 3′ UTR through RNA interference pathways||21–25 nt|
|siRNA||Small interfering RNA||Double-stranded RNA that guides sequence-specific degradation of target mRNA through RNA interference pathway||10–25 bp|
|piRNA||Piwi-interacting RNA||A large class of small ncRNAs involved in retrotransposon silencing through interactions with piwi proteins||26–31 nt|
|Y-RNA||Y RNA||Components of the Ro ribonucleoprotein complex, repressing its activity. It is also required for DNA replication||69–112 nt|
|circRNA||Circular RNA||RNA derived from precursor mRNAs forming covalently closed continuous loop. It is more resistant to exonuclease-mediated degradation||1–5 exons|
|lncRNA||Long noncoding RNA||A non–protein-coding transcript with >200 nt||>200 nt|
Of the various classes of ncRNAs, miRNAs and lncRNAs are the most numerous and are probably the best-known ncRNA groups (numbers of publications concerning miRNAs and lncRNAs are shown in Figure 1). Having different regulatory functions, these types of ncRNAs are important players in the majority of cellular processes, including hematopoiesis, and their abnormal function has serious implications for phenotypes. Deregulation of miRNAs and lncRNAs is frequently found in hematopoietic disorders, contributing substantially to the disease development and progression. Therefore, the following sections of this review will primarily focus on these two categories of ncRNAs and their functions in MDS.
miRNAs are short single-stranded noncoding RNA molecules of approximately 21–25 nt in length. Their sequences are highly conserved in both plants and animals and are thought to be an evolutionarily ancient component of gene regulation. miRNAs posttranscriptionally regulate gene expression through the RNA interference pathway. During the last decade, it has repeatedly been proven that miRNAs play crucial roles in a wide variety of biological processes such as development, differentiation, proliferation, and apoptosis. Because miRNAs influence the expression of genes involved in fundamental signaling pathways, their deregulation often triggers various pathological processes, including cardiovascular diseases, neurological diseases, and cancer.
The first miRNA, lin-4 from
The biogenesis of miRNAs is a multistep process. miRNA genes are transcribed from genomic DNA by RNA polymerase II, resulting in primary miRNA (pri-miRNA) transcripts that usually encode sequences for several miRNAs. In the nucleus, these pri-miRNAs are cleaved by endonuclease Drosha, releasing approximately 70 nucleotide-long hairpin precursor miRNAs (pre-miRNAs). Pre-miRNAs are transported into the cytoplasm, where they are cleaved by Dicer into dsRNA duplexes containing both mature miRNA strand (miRNA) and its complementary strand (miRNA*). In general, the mature miRNA strands are preferentially loaded into the miRNA-induced silencing complex (miRISC), whereas the complementary strands are excluded and degraded. Once processed from the hairpin and loaded into the silencing complex, the miRNA pairs with messenger RNA (mRNA) to direct posttranscriptional repression. At sites with extensive pairing complementarity, the miRNA directs argonaute-catalyzed mRNA cleavage. More commonly, however, the miRNAs direct translational repression, mRNA destabilization, or a combination of the two .
It has been shown that an individual miRNA is able to control the expression of more than one target mRNA and that each mRNA may be regulated by several miRNAs. Generally, it is believed that miRNAs regulate more than 30% of protein-coding genes in the human genome . The ability of miRNAs to interact with thousands of mRNAs has raised intensive interest in their role in physiological and pathological conditions. Like mRNAs, the majority of miRNAs are expressed in tissue-specific manners. For example, miR-122 is preferentially expressed in liver , miR-124 in neurological tissues , miR-133 in muscles , and miR-208a in heart . Moreover, it has been demonstrated that changes in the spectrum of tissue miRNAs correlate with various pathophysiological conditions .
3.1. Extracellular miRNAs
In recent years, cell-free circulating miRNAs have been found in various body fluids such as blood, cerebrospinal fluid, saliva, and urine . The first extracellular small RNAs were observed in blood in 2004 . Unlike the comprehensively described function of cellular miRNAs, the function of miRNAs present in the extracellular environment remains somewhat speculative. However, a growing body of evidence has suggested that these molecules are not mere leftovers of cellular degradation without any specific functions, but that active exchange of miRNAs between cells can play an important role in long-distance cell-to-cell communication.
In 2008, Mitchell et al.  reported that extracellular miRNAs were stable in human plasma/serum. This high stability of circulating miRNAs despite high levels of RNase activity in blood indicates that circulating miRNAs must somehow be protected from degradation. To date, a number of miRNA carriers have been described: membrane-derived vesicles (shedding vesicles, exosomes), lipoproteins, and ribonucleoprotein complexes (with argonaute-2 (AGO2) or nucleophosmin 1 (NPM1) proteins) have been found to transport extracellular miRNAs. It has been reported that the sorting of miRNAs into various types of vesicles can be selective. Diehl et al.  compared the content of miRNAs in microvesicles and their maternal cells. These authors demonstrated a significantly varied spectrum of miRNAs in both samples, suggesting a selective packaging of miRNAs into microvesicles.
3.2. miRNAs in normal hematopoiesis
The differentiation and homeostasis of the hematopoietic system requires complex and interconnected molecular networks that need careful regulation. During the last decade, the role of miRNAs in the hematopoietic system has been extensively studied, and many miRNAs serving as critical regulators of both normal immune functions and diseases have been discovered (Figure 2).
The first study of a role of miRNAs in the differentiation of the immune system showed that forced expression of miR-181 in hematopoietic stem cells (HSCs) markedly increased the number of B lymphocytes, with a concomitant reduction of T lymphocytes . Since then, other miRNAs specific for the maintenance of the “stemness” of HSCs and for the development of individual blood cell lines have been determined. For example, miR-125a was found to be sufficient as a single miRNA to modulate HSC self-renewal and numbers and to protect lineage-negative progenitor cells from apoptosis . A key miRNA that regulates granulocytic differentiation and function is miR-223. This miRNA shows a highly lineage-specific pattern of expression with low levels in HSCs and common myeloid progenitors. The expression of this miRNA is steadily upregulated during differentiation to granulocytes and is repressed during differentiation to the alternative monocytic fate . miR-451 is expressed predominantly in erythroid cells, and its expression is significantly increased during maturation of erythrocytes. In contrast, miR-221 and miR-222 are downregulated during erythroid differentiation. This downregulation enables the expression of their target gene KIT (KIT proto-oncogene receptor tyrosine kinase, CD117), whose activation triggers erythroblast expansion . Another miRNA, miR-150, is highly expressed in mature lymphocytes, whereas it is not active in HSCs. The target gene of miR-150 is transcription factor MYB (myeloblastosis) that regulates lymphocyte development . miR-155, with its high levels in activated B cells, T cells, and monocytes, also participates in lymphoid differentiation. The development of B cells is positively regulated by miRNAs encoded by cluster miR-17-92. This cluster of miRNAs inhibits the expression of the apoptotic gene BIM (proapoptotic BH3-only Bcl-2 family member) and thus plays a key role in pro-B cells to pre-B cells transition . In our laboratory, we analyzed miRNA expression in individual cell types from the peripheral blood of healthy individuals and determined a panel of 13 miRNAs whose expression profile enables differentiation of individual blood cell lines and determination of the cellular origin of in vitro cultured lines .
3.3. miRNAs in malignant hematopoiesis
In oncogenesis, miRNAs act both as oncogenes and as tumor suppressors. Mechanisms of deregulation are similar to those of protein-coding genes (chromosome aberrations, mutations, and epigenetic modifications). In 2002, the loss of two miRNAs (miR-15a and miR-16-1) due to a deletion in the 13q14 region in patients with chronic lymphocytic leukemia was described for the first time as directly associated with malignant disease . In subsequent years, several miRNAs with key roles in the pathogenesis and prognosis of hematological malignancies were detected.
Several publications involved investigations of the role of miRNAs in AML. miRNA expression profiling revealed marked differences in miRNA expression between common cytogenetic subtypes of AML. Jongen-Lavrencic et al.  identified upregulation of several miRNAs (miR-382, miR-134, miR-376a, miR-127, miR-299-5p, and miR-323) in AML patients with t(15;17) and significant downregulation of miRNAs from let-7 family in AML with t(8;21) as well as in AML with inv(16). Moreover, miRNA signatures have been reported to be associated with recurrent molecular abnormalities in cytogenetically normal AML . For example, upregulation of miR-10, let-7, and miR-29 family members and downregulation of miR-204 and miR-128a were found in AML with
3.4. miRNA deregulation in MDS
Several preliminary reports focused on identification of miRNA expression profiles that were either common in MDS or specific for individual MDS subcategories. For example, Pons et al.  measured levels of expression of 25 mature miRNAs in mononuclear cells (MNCs) of MDS patients. The authors reported overexpression of miRNA cluster miR-17-92 in MDS and differential expression of miR-15a and miR-16 between low- and high-risk subgroups of patients. Hussein et al.  showed that the miRNA profiles in BM cells discriminated MDS with chromosomal alterations from patients with normal karyotypes. Sokol et al.  examined miRNA signature in BM MNCs and found deregulation of several miRNAs (increase of miR-222 and miR-10a; decrease of miR-146a, miR-150, and let-7e) in MDS. In our study, we analyzed miRNA expression on a genome-wide level in CD34+ BM cells. We observed significant differences in miRNA expression between early and advanced MDS; an apparent changeover was found between MDS with excess blast 1 (MDS-EB1) and MDS-EB2 subtypes. In particular, we identified strong upregulation of proapoptotic miR-34a in early subtypes of MDS .
Although many studies were conducted regarding miRNA profiling in MDS, there have been very few overlaps among them. This inconsistency may mirror the heterogeneity of the disease but also may possibly be explained by variations between the protocols and platforms used for miRNA detection.
There are several lines of evidence that many miRNAs are deregulated in MDS; however, the functions of miRNAs in MDS pathogenesis remain rather unknown. Identification of target genes of miRNAs in MDS and their functional proofs in both in vitro cell cultures and in vivo animal models are necessary to realizing this goal.
3.4.1. miRNAs in MDS with del(5q)
One of the best-characterized MDS subtypes is MDS with isolated del(5q), formerly referred to as 5q− syndrome. Haploinsufficiency of specific genes within common deleted region (CDR) localized in 5q31.3-5q33 locus is essential for the specific phenotype of MDS with del(5q). In addition to protein-coding genes, 13 genes encoding miRNAs are located in CDR. Most importantly, Starczynowski et al.  correlated del(5q) haploinsufficiency with loss of two miRNAs that are abundant in hematopoietic stem/progenitor cells (HSPCs), miR-145 and miR-146a. Knockdown of miR-145 and miR-146a together in mouse HSPCs resulted in thrombocytosis, mild neutropenia, and megakaryocytic dysplasia .
Kumar et al.  showed that miR-145 loss in MDS with del(5q) affects megakaryocyte and erythroid differentiation. These authors found that miR-145 functions through the repression of FLI1 (Friend Leukemia Integration 1 Transcription Factor), a megakaryocyte and erythroid regulatory transcription factor. Inhibition of miR-145 increases the production of megakaryocytic cells relative to that of erythroid cells. Moreover, the authors proved that combined loss of miR-145 and RPS14 (a ribosomal gene that is required for the maturation of 40S ribosomal subunits and that maps to the CDR) cooperates to alter erythroid-megakaryocytic differentiation in a manner similar to that of the 5q− syndrome .
In our studies, we detected high expression of miR-34a in MDS del(5q) patients [36, 39]. The expression of miR-34a is induced by p53, activating apoptosis through inhibition of BCL2 gene (B-Cell CLL/Lymphoma 2, Apoptosis Regulator). This result is consistent with the increased apoptosis of progenitor cells seen in MDS del(5q).
3.4.2. miRNAs related to prognosis of MDS
Several publications have focused on miRNA expression in MDS with regard to prognostic potential. The earliest study in this area associated miRNA profiles with the International Prognostic Scoring System (IPSS) score . A unique signature consisting of 10 miRNAs was closely associated with IPSS risk category permitting discrimination between lower- and higher-risk disease. Selective overexpression of miR-181 family members was detected in higher risk MDS, indicating pathogenetic overlap with AML. Survival analysis revealed shorter survival in patients with high expression of miR-181 family than in patients with low miR-181 expression .
Another miRNA that has been identified as having prognostic value in MDS is miR-22 . This miRNA was upregulated in MDS and its level correlated with poor survival. Transgenic mice expressing miR-22 in the hematopoietic cells displayed reduced levels of global 5-hydroxymethylcytosine and increased HSC self-renewal accompanied by defective differentiation. Over time, these mice developed MDS. Interestingly, TET2 gene (Ten-Eleven Translocation 2, Tet Methylcytosine Dioxygenase 2) was identified as a key target of miR-22 in this context . TET2 is a major regulator of DNA demethylation by conversion of methylated cytosine into 5-hydroxymethylcytosine.
3.4.3. miRNAs in the treatment of MDS
Expression profiles of miRNAs also appear to be applicable predictors of treatment responses. Lenalidomide is an immunomodulatory and antiangiogenic agent used for the treatment of MDS with del(5q). In recent years, several studies have analyzed miRNA expression levels before and after lenalidomide treatment in these patients [41, 42, 43, 44]. Oliva et al.  investigated expression of selected genes/miRNAs at baseline and after 3 and 6 months of lenalidomide treatment. These authors showed that the expression levels of miR-145, miR-146, and miR-155 gradually increased during the course of the treatment. The significant role of miR-143 and miR-145 in response to lenalidomide was confirmed by Venner et al. , who showed that lenalidomide selectively abrogated progenitor activity in cells depleted of miR-143 and miR-145, supporting their key role in the sensitivity to lenalidomide in MDS with del(5q). In our studies, the most significant changes in expression levels (decreases) after lenalidomide treatment showed miR-34a and several miRNAs clustered within the 14q32 locus [43, 44]. However, a question remains as to whether the changes in expression levels were due to direct response to lenalidomide or whether they were caused by a reduction of the pathological clone.
Therapy with hypomethylating agents (HMAs) such as azacitidine and decitabine is currently considered to be the standard therapy for higher-risk MDS and AML with myelodysplasia-related changes. Several studies investigated miRNA expression with respect to HMA treatment in AML [45, 46, 47]. Blum et al.  proposed miR-29b as a predictive factor for the stratification of older AML patients treated with decitabine; however, this was not confirmed by other studies . Butrym et al.  showed that low expression of miR-181 at diagnosis was a predictor of complete remission and prolonged survival in a subset of older AML patients treated with azacitidine.
In relation to HMA therapy in MDS, it was found that the level of extracellular miR-21 was associated with overall response rate and progression-free survival . Furthermore, reduced expression of miR-124 (caused by abnormal methylation) was found in MDS/AML patients responding to decitabine. These patients exhibited significantly lower expression levels of the CDK6 gene (cyclin-dependent kinase 6) that is the target of miR-124 . Moreover, hypermethylation of miR-124-3p gene appeared to be a good prognostic marker of overall survival . In our report, we found that the overall response rate to azacitidine treatment was significantly higher in MDS/AML patients with upregulated miR-17-3p and downregulated miR-100-5p and miR-133b and that the high level of miR-100-5p was associated with shorter overall survival .
3.4.4. Extracellular miRNAs in MDS
To date, only a few studies have investigated circulating miRNAs in MDS [48, 52, 53, 54]. Two papers [48, 52] focused on specific circulating miRNAs (miR-21, let-7a, and miR-16) that were preselected based on information regarding their deregulation in blood cells and their importance in similar diseases. Researchers monitored the plasma/serum levels of these miRNAs and showed that their levels could serve as prognostic markers for MDS. Kim et al.  showed that serum level of miR-21 was significantly associated with overall response rate and progression-free survival in MDS patients treated with HMAs. The publication that studied let-7a and miR-16 demonstrated that high plasma levels of these miRNAs can serve as semi-invasive markers of poor outcome for MDS patients .
Zuo et al.  measured expression of 800 human miRNAs in MDS plasma. These authors identified a 7-miRNA signature (let-7a, miR-144, miR-16, miR-25, miR-451, miR-651, and miR-655) as an independent predictor of survival in MDS patients with normal karyotypes.
In our study, we investigated the spectrum (2006 human miRNAs) of circulating miRNAs in the plasma of MDS patients . With regard to prognosis, the levels of miR-27a-3p, miR-150-5p, miR-199a-5p, miR-223-3p, and miR-451a were lower in higher risk MDS. Moreover, miR-451a was an independent predictor of progression-free survival, and there was a significant association of miR-223-3p with overall survival .
These pioneer studies suggested that plasma levels of specific miRNAs were associated with MDS patient outcome and may add information beyond the currently used scoring systems. Despite these early promising results, there remain insufficient data regarding the full spectrum of extracellular RNAs in MDS. To date, the possible presence of various forms of small noncoding RNAs (apart from mature miRNAs), pathways for their protection, and identification of their cells of origin have not been explored in MDS. These missing information would expand the knowledge regarding extracellular RNAs in this disease, and beyond that, it would definitely contribute to better interpretation of alterations of individual miRNAs with the potential to become specific prognostic markers in MDS.
4. Long noncoding RNAs
lncRNAs form perhaps the most numerous group of ncRNAs. These RNAs are defined as protein-noncoding transcripts longer than 200 nucleotides. This length was proposed to distinguish lncRNAs from small noncoding RNAs. In contrast with protein-coding RNAs, lncRNAs contain only short open reading frames or completely lack them. This group of ncRNAs is characterized by high levels of structural and functional diversity, low levels of GC nucleotides, and lower expression levels, in contrast with protein-coding transcripts. lncRNAs are transcribed by RNA polymerase II or III and subsequently can be spliced and polyadenylated at the 3′ end or may contain a 5′ cap, depending on their biogenesis. Their expression is developmental and tissue-specific.
Some lncRNAs regulate (negatively or positively) the expression of genes in
According to their position relative to protein-coding mRNAs, lncRNAs are further subcategorized into several groups. Long intergenic noncoding RNAs (lincRNAs) are lncRNAs that are located between annotated protein-coding genes and that are at least 1 kb away from the nearest protein-coding genes. Intronic lncRNAs are coded within introns of protein-coding genes. Sense and antisense lncRNAs are transcribed from the sense or antisense strands of protein-coding genes and often contain exons of this gene with mutual overlap. Bidirectional lncRNAs are oriented head-to-head with protein-coding genes within 1 kb. Bidirectional lncRNA transcripts often exhibit similar expression patterns to those of their protein-coding counterpart, suggesting that they may be subject to shared regulatory pressures. Another group of lncRNA is TERRA (Telomeric Repeat-Containing RNA), transcribed from constitutive heterochromatin-rich regions, or T-UCR (Transcribed Ultraconserved Regions), transcribed from highly conserved regions of the genome.
Although more than 100,000 lncRNAs have been identified to date, only a small number of them have been characterized in detail. To integrate the data describing various lncRNAs, their expression profiles, molecular features, and functions in a variety of cell systems, several databases containing a comprehensive list of lncRNAs have been developed and are continually being updated. Among the most comprehensive databases are LNCipedia (compendium of human lncRNAs, lncipedia.org) , lncRNAdb (reference database for functional lncRNAs, lncRNAdb.org) , and NRED (database of lncRNA expression, nred.matticklab.com) .
4.1. lncRNAs in normal hematopoiesis
Recently, lncRNAs have emerged as important regulators of cell fate. These RNAs play a variety of roles in controlling various steps in hematopoietic differentiation, including maintenance of HSCs and differentiation of myeloid, erythroid, and lymphoid lineages. To date, there have been several descriptions of lncRNAs crucial for correct function of the hematopoietic system (Table 2). The first hematopoiesis-associated lncRNA, EGOT (Eosinophil Granule Ontogeny Transcript), was described in 2007. EGOT, a conserved transcript localized antisense to ITPR1 (Inositol 1,4,5-Trisphosphate Receptor Type 1) that modulates the development of eosinophils, is normally expressed in human CD34+ HSCs, and its expression level increases during eosinophil development, helping to regulate production of eosinophil granule proteins .
(Eosinophil Granule Ontogeny Transcript)
|3p26.1, antisense to ITPR1 gene||Modulates development of eosinophils|||
|H19||11p15.5, antisense to IGF2||Maintains quiescence of adult HSCs|||
(Hox Antisense Intergenic RNA Myeloid 1)
|7p15.2, encoded within the HOXA gene cluster||Induces differentiation of myeloid cells|||
|lincRNA-EPS||Mouse 4qC7||Promotes red blood cell maturation|||
|T-ALL-R-LncR1||6q24.3||Induces apoptosis in T-ALL|||
(Leukemia-Induced Noncoding Activator RNA)
|15q26.3||Positive regulator of cell division|||
(Hox Transcript Antisense RNA)
|12q13.13, encoded within the HOXC gene cluster||Oncogene, promotes chromatin relocalization|||
(Metastasis Associated Lung Adenocarcinoma Transcript)
|11q13.1||Regulates transcription and cell cycle|||
(Maternally Expressed Gene 3)
(X-Inactive Specific Transcript)
|Xq13.2||Regulates X chromosome inactivation during embryogenesis|||
The relatively well-known lncRNA, H19, maintains quiescence of adult HSCs. The H19 transcript was in fact the first lncRNA to be identified, enriched in the embryonic fetal liver but downregulated after birth . H19 is active in long-term HSCs and becomes gradually downregulated in short-term HSCs and multipotent progenitors. Deletion of H19 from the maternal allele resulted in increased HSC activation and proliferation as well as impaired repopulating ability. This effect is mediated by derepression of maternal IGF2 (Insulin-Like Growth Factor 2) expression and by increased IGF1R (IGF1 Receptor) translation, resulting in increased signaling through the IGF1R .
Zhang et al.  identified HOTAIRM1 (Homeobox Antisense Intergenic RNA Myeloid 1), which is encoded within the human HOXA (Homeobox A) gene cluster and plays a role in the differentiation of myeloid cells. The expression of this transcript is upregulated during granulocyte differentiation. Knockdown of HOTAIRM1 reduced transcription of HOXA1 and HOXA4 in an acute promyelocytic leukemia cell line, resulting in a decreased expression of genes associated with granulocyte activation, defense response, and maturation .
Hu et al.  studied the lncRNA transcriptome of the erythroid lineage and uncovered numerous erythroid-specific lncRNAs that become induced during terminal differentiation of mouse fetal liver red blood cells in vivo. These authors showed that lincRNA-EPS (erythroid prosurvival) acts to promote red blood cell maturation by downregulating proapoptotic pathways. Knockdown of lincRNA-EPS severely compromised terminal differentiation of erythroid progenitors and resulted in elevated apoptosis. Conversely, its ectopic expression protected erythroid progenitors from apoptosis triggered by erythropoietin starvation. Functional studies indicated that lincRNA-EPS acts by repressing a number of proapoptotic proteins, most prominently the caspase-activating adaptor protein Pycard .
Recent studies have also provided evidence for the importance of several lncRNAs in immune cell function. For example, lncRNA NeST (Nettoie Salmonella pas Theiler’s, cleanup Salmonella not Theiler’s), also named Tmevpg1, modulates the ability of mice to respond to viral and bacterial infections. NeST is specifically expressed by the TH1 subset of helper T cells. The expression of NeST regulates the degree of inflammation induced by infecting pathogens, such as Thelier’s virus or Salmonella . lncRNA-Cox2 (cyclooxygenase 2) acts during inflammatory signaling by modulating the expression of several immune response genes via interactions with regulatory complexes .
4.2. lncRNAs in malignant hematopoiesis
lncRNAs not only participate in normal hematopoiesis but also contribute to the pathogenesis of hematologic malignancies, representing a new class of potential biomarkers and therapeutic targets. These RNAs have significantly different expression levels in primary tumors and metastases, functioning both as oncogenes or as tumor suppressors. Some cancer-related lncRNAs could affect the development and progression of tumor by means of p53, polycomb repressive complex 2 (PRC2), and other signaling pathways. Others are not observed in normal tissue but are detected in cancer. For example, T-ALL-R-lncR1 appears to induce (together with protease-activated receptor 4 (PAR-4)) cellular apoptosis in T cell acute lymphoblastic leukemia cells (T-ALL) . LUNAR1 (leukemia-induced noncoding activator RNA) is highly expressed in T-ALL cells, and its expression is dependent on signaling through the oncogenic NOTCH1 receptor . lncRNAs HOTAIR (homeobox transcript antisense RNA) and MALAT-1 (metastasis-associated lung adenocarcinoma transcript) are associated with metastasis and recurrence [68, 69].
4.2.1. lncRNA deregulation in MDS
The first study describing lncRNA in the context of MDS was published in 2010 by Benetatos et al.  who studied lncRNA MEG3 (maternally expressed gene 3). Abnormal methylation of its promoter was observed in a third of MDS patients and in half of AML patients . MEG3 was the first lncRNA described to have a tumor suppressor function. MEG3 is expressed in many human normal tissues, and numerous studies have demonstrated that its expression level is lost in various cancers. Low expression of MEG3 is associated with an increased risk of metastasis and poor prognosis in cancer patients [71, 72].
In 2013, Yildirim et al.  conditionally deleted lncRNA XIST (X-inactive specific transcript) in mice hematopoietic cells. XIST is perhaps the most well-understood lncRNA to date. This lncRNA is located on the X chromosome and is required for X chromosome inactivation during embryogenesis. Yildirim et al.  demonstrated that mutant females developed a highly aggressive myeloproliferative neoplasm and MDS (mixed MPN/MDS) with 100% penetrance.
The first study examining the deregulation of lncRNAs on a genome-wide level in MDS was published in 2017 . The authors combined NGS and microarray data in CD34+ BM cells and identified several lncRNAs (linc-ARFIP1-4, linc-TAAR9-1, lincC2orf85, linc-RNFT2-1, and linc-RPIA) deregulated in MDS-EB2. In the same year, Yao et al.  profiled lncRNA expressions in 176 adult patients with primary MDS and identified four lncRNAs (TC07000551.hg.1, TC08000489.hg.1, TC02004770.hg.1, and TC03000701) with expression levels significantly associated with overall survival. Subsequently, the authors constructed a risk-scoring system with the weighted sum of these four lncRNAs. Higher lncRNA scores were associated with higher marrow blast percentages, higher-risk subtypes of MDS, complex cytogenetic changes, and mutations in RUNX1, ASXL1, TP53, SRSF2, and ZRSR2, whereas they were inversely correlated with the SF3B1 mutation. Patients with higher lncRNA scores had significantly shorter overall survival and higher 5-year leukemic transformation rate than did those with lower scores .
Although increasing numbers of deregulated lncRNAs are currently being described in MDS, only a few have been functionally characterized so far. Transcriptomic data may be used to construct network modules consisting of lncRNAs and protein-coding genes to enable functional analysis of lncRNAs with unknown functions. These networks are subsequently linked with annotated signaling pathways and gene ontologies. The resulting outputs provide a degree of functional annotation for differentially expressed lncRNAs in the disease and their potential roles in pathophysiology .
5. Other types of noncoding RNAs in MDS
Only very limited information regarding other groups of ncRNAs have been published for MDS to date. However, we can anticipate that introduction of next-generation sequencing of RNAs (so-called RNA-seq) will bring to MDS research many novel insights regarding various ncRNAs in the near future. This technology enables sensitive global detection of various RNAs across an unparalleled dynamic range. Particularly, small RNA-seq is predominantly used for detection of miRNAs. However, during library preparation, small RNAs are selected by electrophoresis with sizes typically ranging from 20 to 50 nt. This range of size selection allows for the capture of many other species of small RNAs in addition to miRNAs.
5.1. Piwi-interacting RNAs
In 2011, Beck et al.  conducted one of the early studies to apply small RNA-seq in MDS. These authors compared expression of small RNAs between low-grade (refractory anemia, RA) and high-grade (MDS-EB2) MDS patients and demonstrated the first evidence of piwi (P-element-Induced Wimpy Testis)-interacting RNAs (piRNAs) in MDS BM cells and their particular enrichment in low-grade MDS. PiRNAs are a relatively newly defined class of small ncRNAs with lengths from 26 to 32 nt. These RNAs lack sequence conservation and are more complex than miRNAs. PiRNAs have been linked to both epigenetic and posttranscriptional gene silencing of retrotransposons and other genetic elements in germline cells, particularly those involved in spermatogenesis.
Transcription of particular piwi proteins (
5.2. Transfer RNAs
The abovementioned pioneer study from Beck et al.  also provided an early insight into the deregulation of tRNAs in MDS. The authors showed that ratios of tRNA to rRNA were significantly higher in MDS-EB2 compared to those of RA and controls. Because tRNAs are building blocks for protein synthesis and are required during translation, this may indicate an increased regulation of translation at this disease stage. Interestingly, a significant increase of tRNAs in tumor samples was reported by Pavon-Eternod et al.  In addition, tRNAs have been shown to inhibit cytochrome c-activated apoptosis . Taken together, Beck et al.  hypothesized that high tRNA content seen in EB2 may contribute to the two well-known characteristics of high-grade MDS, decreased apoptosis, and high rate of leukemic transformation.
Guo et al.  performed small RNA-seq in paired pre- and posttreatment samples from MDS patients receiving therapy with HMAs. In the sequencing data, the number of reads aligned to tRNA-derived small RNAs (tDRs) (78.81%) vastly outnumbered those aligning to miRNAs (4.43% of reads). The tRNA fragments that were captured by miRNA-seq might be a result of either active cleavage or artifacts of the miRNA-seq library construction. The authors identified six tDR fragments that were differentially expressed between MDS and normal samples. Three tDRs demonstrated increased expression in MDS (chrM.tRNA10.TC, chr12.tRNA8.AlaTGC, and chr16.tRNA4.ProAGG), while three were decreased (chr1.tRNA58-LeuCAA, chr19.tRNA8-SeC(e)TCA (SeC(e)TCA), and chr19.tRNA4-ThrAGT). Moreover, they identified a panel of four tRNA fragments (chr6.tRNA157.ValCAC, chr11.tRNA17.ValTAC, chrM.tRNA12.TS1, and chrX.tRNA4.ValTAC), whose combined expression in the pretreatment samples together was predictive of the likelihood of response. Deeper focus on mitochondrial tRNAs revealed that MT-TS1 (mitochondrially encoded tRNA serine 1) was the only mitochondrial tRNA to have a significant association with treatment response .
The discovery of ncRNAs has initiated a new era in molecular biology, completely changing our view of “junk” DNA that it is no longer considered unnecessary ballast. Mouse models have clearly demonstrated key functions of ncRNAs in regulatory networks and their ability to significantly influence biological processes. In the hematopoietic system, ncRNAs represent important regulators of HSC “stemness” and differentiation; therefore, it is not surprising that deregulation of ncRNAs also occurs in MDS.
Currently, we possess comprehensive information regarding the impact of miRNA deregulation on the pathogenesis of MDS. The efforts of current research activities aim to apply these findings to clinical practice, testing the potential diagnostic/prognostic value of selected miRNAs for MDS. However, miRNAs also represent promising therapeutic agents or targets. miRNA-based drugs are designed to reduce the expression of oncogenic miRNAs or, conversely, to increase levels of miRNAs with tumor suppressor functions. Unlike targeted inhibition or activation of a single protein-coding gene, the administration of miRNA antagonists or their mimics may potentially improve the desired effects, as these molecules can regulate several genes, often in specific signaling pathways implicated in tumorigenesis. Several pharmaceutical companies already have miRNA therapeutics in their developmental pipelines . In 2012, the first cancer-targeted miRNA drug, MRX34 (a liposome-based miR-34 mimic), entered phase I clinical trials in patients with advanced hepatocellular carcinoma, and this mimic has attracted considerable attention from both academic researchers and pharmaceutical companies . MRG-106, a synthetic antagonist of miRNA-155, is currently being tested by MIRagen Therapeutics in patients with cutaneous T-cell lymphoma . However, testing miRNAs as potential therapeutic agents or targets in MDS therapy still requires initial exploration in in vitro models before evolving to future clinical trials.
Information regarding the contribution of other categories of ncRNAs, including lncRNAs, to the pathogenesis of MDS remains scarce. However, given the large number of ncRNAs encoded in the human genome and the complexity of their interactions, it can be expected that, in the near future, we will reveal a number of ncRNAs involved in MDS. It can also be anticipated that we will identify new predictive markers of progression and responses to therapy among these molecules.
To conclude, the diagnostic and therapeutic possibilities of ncRNAs undoubtedly have profound potential in MDS. However, although the effects of some miRNAs have already been demonstrated, it is certain that the importance of ncRNAs in MDS will be fully understood only in the future and that many years of research and clinical trials remain before the eventual application of ncRNAs in clinical practice to classify, monitor, and treat this disease.
This work was supported by the research grants no. 16-33617A and 17-31398A and the project for conceptual development of research organization no. 00023736 (all from the Ministry of Health of the Czech Republic).
Conflict of interests
The authors declare that they have no competing interests.
Garcia-Manero G. Myelodysplastic syndromes: 2014 update on diagnosis, risk-stratification, and management. American Journal of Hematology. 2014; 89:97-108. DOI: 10.1002/ajh.23642
Bejar R, Stevenson K, Abdel-Wahab O, Galili N, Nilsson B, Garcia-Manero G, et al. Clinical effect of point mutations in myelodysplastic syndromes. The New England Journal of Medicine. 2011; 364:2496-2506. DOI: 10.1056/NEJMoa1013343
Papaemmanuil E, Gerstung M, Malcovati L, Tauro S, Gundem G, Van Loo P, et al. Clinical and biological implications of driver mutations in myelodysplastic syndromes. Blood. 2013; 122:3616-3627. DOI: 10.1182/blood-2013-08-518886
Lee RC, Feinbaum RL, Ambros V. The C. elegansheterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14. Cell. 1993; 75:843-854
Wightman B, Ha I, Ruvkun G. Posttranscriptional regulation of the heterochronic gene lin-14 by lin-4 mediates temporal pattern formation in C. elegans. Cell. 1993; 75:855-862
Pennisi E. Genomics. ENCODE project writes eulogy for junk DNA. Science. 2012; 337:1159-1161. DOI: 10.1126/science.337.6099.1159
ENCODE Project Consortium. An integrated encyclopedia of DNA elements in the human genome. Nature. 2012; 489:57-74. DOI: 10.1038/nature11247
Palazzo AF, Lee ES. Non-coding RNA: What is functional and what is junk? Frontiers in Genetics. 2015; 6:2. DOI: 10.3389/fgene.2015.00002
Dozmorov MG, Giles CB, Koelsch KA, Wren JD. Systematic classification of non-coding RNAs by epigenomic similarity. BMC Bioinformatics. 2013; 14(Suppl 14):S2. DOI: 10.1186/1471-2105-14-S14-S
Bartel DP. MicroRNA target recognition and regulatory functions. Cell. 2009; 136:215-233. DOI: 10.1016/j.cell.2009.01.002
Lewis BP, Burge CB, Bartel DP. Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets. Cell. 2005; 120:15-20. DOI: 10.1016/j.cell.2004.12.035
Lewis AP, Jopling CL. Regulation and biological function of the liver-specific miR-122. Biochemical Society Transactions. 2010; 38:1553-1557. DOI: 10.1042/BST0381553
Sun AX, Crabtree GR, Yoo AS. MicroRNAs: Regulators of neuronal fate. Current Opinion in Cell Biology. 2013; 25:215-221. DOI: 10.1016/j.ceb.2012.12.007
Luo W, Nie Q, Zhang X. MicroRNAs involved in skeletal muscle differentiation. Journal of Genetics and Genomics. 2013; 40:107-116. DOI: 10.1016/j.jgg.2013.02.002
van Rooij E, Sutherland LB, Qi X, Richardson JA, Hill J, Olson EN. Control of stress-dependent cardiac growth and gene expression by a microRNA. Science. 2007; 316:575-579. DOI: 10.1126/science.1139089
Lu J, Getz G, Miska EA, Alvarez-Saavedra E, Lamb J, Peck D, et al. MicroRNA expression profiles classify human cancers. Nature. 2005; 435:834-838. DOI: 10.1038/nature03702
Weber JA, Baxter DH, Zhang S, Huang DY, Huang KH, Lee MJ, et al. The microRNA spectrum in 12 body fluids. Clinical Chemistry. 2010; 56:1733-1741. DOI: 10.1373/clinchem.2010.147405
El-Hefnawy T, Raja S, Kelly L, Bigbee WL, Kirkwood JM, Luketich JD, et al. Characterization of amplifiable, circulating RNA in plasma and its potential as a tool for cancer diagnostics. Clinical Chemistry. 2004; 50:564-573. DOI: 10.1373/clinchem.2003.028506
Mitchell PS, Parkin RK, Kroh EM, Fritz BR, Wyman SK, Pogosova-Agadjanyan EL, et al. Circulating microRNAs as stable blood-based markers for cancer detection. Proceedings of the National Academy of Sciences of the United States of America. 2008; 105:10513-10518. DOI: 10.1073/pnas.0804549105
Diehl P, Fricke A, Sander L, Stamm J, Bassler N, Htun N, et al. Microparticles: Major transport vehicles for distinct microRNAs in circulation. Cardiovascular Research. 2012; 93:633-644. DOI: 10.1093/cvr/cvs007
Chen CZ, Li L, Lodish HF, Bartel DP. MicroRNAs modulate hematopoietic lineage differentiation. Science. 2004; 303:83-86. DOI: 10.1126/science.1091903
Guo S, Lu J, Schlanger R, Zhang H, Wang JY, Fox MC, et al. MicroRNA miR-125a controls hematopoietic stem cell number. Proceedings of the National Academy of Sciences of the United States of America. 2010; 107:14229-14234. DOI: 10.1073/pnas.0913574107
Johnnidis JB, Harris MH, Wheeler RT, Stehling-Sun S, Lam MH, Kirak O, et al. Regulation of progenitor cell proliferation and granulocyte function by microRNA-223. Nature. 2008; 451:1125-1129. DOI: 10.1038/nature06607
Bruchova H, Yoon D, Agarwal AM, Mendell J, Prchal JT. Regulated expression of microRNAs in normal and polycythemia vera erythropoiesis. Experimental Hematology. 2007; 35:1657-1667. DOI: 10.1016/j.exphem.2007.08.021
Morris VA, Zhang A, Yang T, Stirewalt DL, Ramamurthy R, Meshinchi S, et al. MicroRNA-150 expression induces myeloid differentiation of human acute leukemia cells and normal hematopoietic progenitors. PLoS One. 2013; 8(9):e75815. DOI: 10.1371/journal.pone.0075815
Vasilatou D, Papageorgiou S, Pappa V, Papageorgiou E, Dervenoulas J. The role of microRNAs in normal and malignant hematopoiesis. European Journal of Haematology. 2010; 84:1-16. DOI: 10.1111/j.1600-0609.2009.01348.x
Merkerova M, Belickova M, Bruchova H. Differential expression of microRNAs in hematopoietic cell lineages. European Journal of Haematology. 2008; 81:304-310. DOI: 10.1111/j.1600-0609.2008.01111.x
Calin GA, Dumitru CD, Shimizu M, Bichi R, Zupo S, Noch E, et al. Frequent deletions and down-regulation of micro-RNA genes miR15 and miR16 at 13q14 in chronic lymphocytic leukemia. Proceedings of the National Academy of Sciences of the United States of America. 2002; 99:15524-15529. DOI: 10.1073/pnas.242606799
Jongen-Lavrencic M, Sun SM, Dijkstra MK, Valk PJ, Löwenberg B. MicroRNA expression profiling in relation to the genetic heterogeneity of acute myeloid leukemia. Blood. 2008; 111:5078-5085. DOI: 10.1182/blood-2008-01-133355
Zhao H, Wang D, Du W, Gu D, Yang R. MicroRNA and leukemia: Tiny molecule, great function. Critical Reviews in Oncology/Hematology. 2010; 74:149-155. DOI: 10.1016/j.critrevonc.2009.05.001
Marcucci G, Maharry K, Radmacher MD, Mrózek K, Vukosavljevic T, Paschka P, et al. Prognostic significance of, gene and microRNA expression signatures associated with, CEBPA mutations in cytogenetically normal acute myeloid leukemia with high-risk molecular features: A Cancer and Leukemia Group B Study. Journal of Clinical Oncology. 2008; 26:5078-5087. DOI: 10.1200/JCO.2008.17.5554
Garzon R, Volinia S, Liu CG, Fernandez-Cymering C, Palumbo T, Pichiorri F, et al. MicroRNA signatures associated with cytogenetics and prognosis in acute myeloid leukemia. Blood. 2008; 111:3183-3189. DOI: 10.1182/blood-2007-07-098749
Pons A, Nomdedeu B, Navarro A, Gaya A, Gel B, Diaz T, et al. Hematopoiesis-related microRNA expression in myelodysplastic syndromes. Leukemia & Lymphoma. 2009; 50:1854-1859. DOI: 10.3109/10428190903147645
Hussein K, Theophile K, Büsche G, Schlegelberger B, Göhring G, Kreipe H, et al. Aberrant microRNA expression pattern in myelodysplastic bone marrow cells. Leukemia Research. 2010; 34:1169-1174. DOI: 10.1016/j.leukres.2010.04.012
Sokol L, Caceres G, Volinia S, Alder H, Nuovo GJ, Liu CG, et al. Identification of a risk dependent microRNA expression signature in myelodysplastic syndromes. British Journal of Haematology. 2011; 153:24-32. DOI: 10.1111/j.1365-2141.2011.08581.x
Dostalova Merkerova M, Krejcik Z, Votavova H, Belickova M, Vasikova A, Cermak J. Distinctive microRNA expression profiles in CD34+ bone marrow cells from patients with myelodysplastic syndrome. European Journal of Human Genetics. 2011; 19:313-319. DOI: 10.1038/ejhg.2010.209
Starczynowski DT, Kuchenbauer F, Argiropoulos B, Sung S, Morin R, Muranyi A, et al. Identification of miR-145 and miR-146a as mediators of the 5q− syndrome phenotype. Nature Medicine. 2010; 16:49-58. DOI: 10.1038/nm.2054
Kumar MS, Narla A, Nonami A, Mullally A, Dimitrova N, Ball B, et al. Coordinate loss of a microRNA and protein-coding gene cooperate in the pathogenesis of 5q− syndrome. Blood. 2011; 118(17):4666-4673. DOI: 10.1182/blood-2010-12-324715
Votavova H, Grmanova M, Dostalova Merkerova M, Belickova M, Vasikova A, Neuwirtova R, et al. Differential expression of microRNAs in CD34+ cells of 5q− syndrome. Journal of Hematology & Oncology. 2011; 4:1. DOI: 10.1186/1756-8722-4-1
Song SJ, Ito K, Ala U, Kats L, Webster K, Sun SM, et al. The oncogenic microRNA miR-22 targets the TET2 tumor suppressor to promote hematopoietic stem cell self-renewal and transformation. Cell Stem Cell. 2013; 13:87-101. DOI: 10.1016/j.stem.2013.06.003
Oliva EN, Cuzzola M, Aloe Spiriti MA, Poloni A, Laganà C, Rigolino C, et al. Biological activity of lenalidomide in myelodysplastic syndromes with del5q: Results of gene expression profiling from a multicenter phase II study. Annals of Hematology. 2013; 92:25-32. DOI: 10.1007/s00277-012-1569-0
Venner CP, Woltosz JW, Nevill TJ, Deeg HJ, Caceres G, Platzbecker U, et al. Correlation of clinical response and response duration with miR-145 induction by lenalidomide in CD34(+) cells from patients with del(5q) myelodysplastic syndrome. Haematologica. 2013; 98:409-413. DOI: 10.3324/haematol.2012.066068
Merkerova MD, Krejcik Z, Belickova M, Hrustincova A, Klema J, Stara E, et al. Genome-wide miRNA profiling in myelodysplastic syndrome with del(5q) treated with lenalidomide. European Journal of Haematology. 2015; 95:35-43. DOI: 10.1111/ejh.12458
Krejčík Z, Beličková M, Hruštincová A, Kléma J, Zemanová Z, Michalová K, et al. Aberrant expression of the microRNA cluster in 14q32 is associated with del(5q) myelodysplastic syndrome and lenalidomide treatment. Cancer Genetics. 2015; 208:156-161. DOI: 10.1016/j.cancergen.2015.03.003
Blum W, Garzon R, Klisovic RB, Schwind S, Walker A, Geyer S, et al. Clinical response and miR-29b predictive significance in older AML patients treated with a 10-day schedule of decitabine. Proceedings of the National Academy of Sciences of the United States of America. 2010; 107:7473-7478. DOI: 10.1073/pnas.1002650107
Yang H, Fang Z, Wei Y, Hu Y, Calin GA, Kantarjian HM, et al. Levels of miR-29b do not predict for response in patients with acute myelogenous leukemia treated with the combination of 5-azacytidine, valproic acid, and ATRA. American Journal of Hematology. 2011; 86:237-238. DOI: 10.1002/ajh.21937
Butrym A, Rybka J, Baczyńska D, Poręba R, Mazur G, Kuliczkowski K. Expression of microRNA-181 determines response to treatment with azacitidine and predicts survival in elderly patients with acute myeloid leukaemia. Oncology Letters. 2016; 12:2296-2300. DOI: 10.3892/ol.2016.4970
Kim Y, Cheong JW, Kim YK, Eom JI, Jeung HK, Kim SJ, et al. Serum microRNA-21 as a potential biomarker for response to hypomethylating agents in myelodysplastic syndromes. PLoS One. 2014; 9:e86933. DOI: 10.1371/journal.pone.0086933
Liu H, Pattie P, Chandrasekara S, Spencer A, Dear AE. Epigenetic regulation of miRNA-124 and multiple downstream targets is associated with treatment response in myeloid malignancies. Oncology Letters. 2016; 12(3):2175-2180. DOI: 10.3892/ol.2016.4912
Wang H, Zhang TT, Jin S, Liu H, Zhang X, Ruan CG, et al. Pyrosequencing quantified methylation level of miR-124 predicts shorter survival for patients with myelodysplastic syndrome. Clinical Epigenetics. 2017; 9:91. DOI: 10.1186/s13148-017-0388-5
Krejcik Z, Belickova M, Hrustincova A, Votavova H, Jonasova A, Cermak J, et al. MicroRNA profiles as predictive markers of response to azacitidine therapy in myelodysplastic syndromes and acute myeloid leukemia. Cancer Biomarkers. 2018; 22:101-110. DOI: 10.3233/CBM-171029
Zuo Z, Calin GA, de Paula HM, Medeiros LJ, Fernandez MH, Shimizu M, et al. Circulating microRNAs let-7a and miR-16 predict progression-free survival and overall survival in patients with myelodysplastic syndrome. Blood. 2011; 118:413-415. DOI: 10.1182/blood-2011-01-330704
Zuo Z, Maiti S, Hu S, Loghavi S, Calin GA, Garcia-Manero G, et al. Plasma circulating-microRNA profiles are useful for assessing prognosis in patients with cytogenetically normal myelodysplastic syndromes. Modern Pathology. 2015; 28:373-382. DOI: 10.1038/modpathol.2014.108
Dostalova Merkerova M, Hrustincova A, Krejcik Z, Votavova H, Ratajova E, Cermak J, et al. Microarray profiling defines circulating microRNAs associated with myelodysplastic syndromes. Neoplasma. 2017; 64:571-578. DOI: 10.4149/neo_2017_411
Shi X, Sun M, Liu H, Yao Y, Song Y. Long non-coding RNAs: A new frontier in the study of human diseases. Cancer Letters. 2013; 339:159-166. DOI: 10.1016/j.canlet.2013.06.013
Volders PJ, Verheggen K, Menschaert G, Vandepoele K, Martens L, Vandesompele J, et al. An update on LNCipedia: A database for annotated human lncRNA sequences. Nucleic Acids Research. 2015; 43:4363-4364. DOI: 10.1093/nar/gkv295
Quek XC, Thomson DW, Maag JL, Bartonicek N, Signal B, Clark MB, et al. lncRNAdb v2.0: Expanding the reference database for functional long noncoding RNAs. Nucleic Acids Research. 2015; 43:168-173. DOI: 10.1093/nar/gku988
Dinger ME, Pang KC, Mercer TR, Crowe ML, Grimmond SM, Mattick JS. NRED: A database of long noncoding RNA expression. Nucleic Acids Research. 2009; 37:122-126. DOI: 10.1093/nar/gkn617
Wagner LA, Christensen CJ, Dunn DM, Spangrude GJ, Georgelas A, Kelley L, et al. EGO, a novel, noncoding RNA gene, regulates eosinophil granule protein transcript expression. Blood. 2007; 109:5191-5198. DOI: 10.1182/blood-2006-06-027987
Brannan CI, Dees EC, Ingram RS, Tilghman SM. The product of the H19 gene may function as an RNA. Molecular and Cellular Biology. 1990; 10:28-36
Venkatraman A, He XC, Thorvaldsen JL, Sugimura R, Perry JM, Tao F, et al. Maternal imprinting at the H19-Igf2 locus maintains adult haematopoietic stem cell quiescence. Nature. 2013; 500:345-349. DOI: 10.1038/nature12303
Zhang X, Lian Z, Padden C, Gerstein MB, Rozowsky J, Snyder M, et al. A myelopoiesis-associated regulatory intergenic noncoding RNA transcript within the human HOXA cluster. Blood. 2009; 113:2526-2534. DOI: 10.1182/blood-2008-06-162164
Hu W, Yuan B, Flygare J, Lodish HF. Long noncoding RNA-mediated anti-apoptotic activity in murine erythroid terminal differentiation. Genes & Development. 2011; 25:2573-2578. DOI: 10.1101/gad.178780.111
Gomez JA, Wapinski OL, Yang YW, Bureau JF, Gopinath S, Monack DM, et al. The NeST long ncRNA controls microbial susceptibility and epigenetic activation of the interferon-gamma locus. Cell. 2013; 152:743-754. DOI: 10.1016/j.cell.2013.01.015
Carpenter S, Aiello D, Atianand MK, Ricci EP, Gandhi P, Hall LL, et al. A long noncoding RNA mediates both activation and repression of immune response genes. Science. 2013; 341:789-792. DOI: 10.1126/science.1240925
Zhang L, Xu HG, Lu C. A novel long non-coding RNA T-ALL-R-LncR1 knockdown and Par-4 cooperate to induce cellular apoptosis in T-cell acute lymphoblastic leukemia cells. Leukemia & Lymphoma. 2014; 55:1373-1382. DOI: 10.3109/10428194.2013.829574
Trimarchi T, Bilal E, Ntziachristos P, Fabbri G, Dalla-Favera R, Tsirigos A, et al. Genome-wide mapping and characterization of Notch-regulated long noncoding RNAs in acute leukemia. Cell. 2014; 158:593-606. DOI: 10.1016/j.cell.2014.05.049
Gupta RA, Shah N, Wang KC, Kim J, Horlings HM, Wong DJ, et al. Long non-coding RNA HOTAIR reprograms chromatin state to promote cancer metastasis. Nature. 2010; 464:1071-1076. DOI: 10.1038/nature08975
Zhu L, Liu J, Ma S, Zhang S. Long noncoding RNA MALAT-1 can predict metastasis and a poor prognosis: A meta-analysis. Pathology Oncology Research. 2015; 21:1259-1264. DOI: 10.1007/s12253-015-9960-5
Benetatos L, Hatzimichael E, Dasoula A, Dranitsaris G, Tsiara S, Syrrou M, et al. CpG methylation analysis of the MEG3 and SNRPN imprinted genes in acute myeloid leukemia and myelodysplastic syndromes. Leukemia Research. 2010; 34:148-153. DOI: 10.1016/j.leukres.2009.06.019
Zhang Z, Liu T, Wang K, Qu X, Pang Z, Liu S, et al. Down-regulation of long non-coding RNA MEG3 indicates an unfavorable prognosis in non-small cell lung cancer: Evidence from the GEO database. Gene. 2017; 630:49-58. DOI: 10.1016/j.gene.2017.08.001
Tian ZZ, Guo XJ, Zhao YM, Fang Y. Decreased expression of long non-coding RNA MEG3 acts as a potential predictor biomarker in progression and poor prognosis of osteosarcoma. International Journal of Clinical and Experimental Pathology. 2015; 8:15138-15142
Yildirim E, Kirby JE, Brown DE, Mercier FE, Sadreyev RI, Scadden DT, et al. Xist RNA is a potent suppressor of hematologic cancer in mice. Cell. 2013; 152:727-742. DOI: 10.1016/j.cell.2013.01.034
Liu K, Beck D, Thoms JAI, Liu L, Zhao W, Pimanda JE, et al. Annotating function to differentially expressed LincRNAs in myelodysplastic syndrome using a network-based method. Bioinformatics. 2017; 33:2622-2630. DOI: 10.1093/bioinformatics/btx280
Yao CY, Chen CH, Huang HH, Hou HA, Lin CC, Tseng MH, et al. A 4-lncRNA scoring system for prognostication of adult myelodysplastic syndromes. Blood Advances. 2017; 1:1505-1516. DOI: 10.1182/bloodadvances.2017008284
Beck D, Ayers S, Wen J, Brandl MB, Pham TD, Webb P, et al. Integrative analysis of next generation sequencing for small non-coding RNAs and transcriptional regulation in myelodysplastic syndromes. BMC Medical Genomics. 2011; 4:19. DOI: 10.1186/1755-8794-4-19
Klattenhoff C, Theurkauf W. Biogenesis and germline functions of piRNAs. Development. 2008; 135:3-9. DOI: 10.1242/dev.006486
Pavon-Eternod M, Gomes S, Geslain R, Dai Q, Rosner MR, Pan T. tRNA over-expression in breast cancer and functional consequences. Nucleic Acids Research. 2009; 37:7268-7280. DOI: 10.1093/nar/gkp787
van Raam BJ, Salvesen GS. Transferring death: A role for tRNA in apoptosis regulation. Molecular Cell. 2010; 37:591-592. DOI: 10.1016/j.molcel.2010.02.001
Guo Y, Bosompem A, Mohan S, Erdogan B, Ye F, Vickers KC, et al. Transfer RNA detection by small RNA deep sequencing and disease association with myelodysplastic syndromes. BMC Genomics. 2015; 16:727. DOI: 10.1186/s12864-015-1929-y
Shah MY, Ferrajoli A, Sood AK, Lopez-Berestein G, Calin GA. microRNA therapeutics in cancer—An emerging concept. eBioMedicine. 2016; 12:34-42. DOI: 10.1016/j.ebiom.2016.09.017
Bader AG. miR-34—A microRNA replacement therapy is headed to the clinic. Frontiers in Genetics. 2012; 3:120. DOI: 10.3389/fgene.2012.00120
Foss FM, Querfeld C, Porcu P, Kim YH, Pacheco T, Halwani AS. Phase 1 trial evaluating MRG-106, a synthetic inhibitor of microRNA-155, in patients with cutaneous t-cell lymphoma (CTCL). Journal of Clinical Oncology. 2017; 35(15_suppl):7564-7564. DOI: 10.1200/JCO.2017.35.15_suppl.7564
- These authors contributed equally to this work.