Acute myeloid leukemia(AML) is the most common type of acute leukemia in adults. According to the WHO classification,patients with recurring cytogenetic abnormalities,such as t(8,21), inv(16)(p13q22) or t(15,17) often had a better prognosis.But for those with normal karyotype,there is an another story. Actually, CN-AML is a heterogeneous disease. Although with the advent of high-throughput sequencing and other methods, many genetic changes (such as mutations of gene FLT3,NPM1) which were closely related to the prognosis of CN-AML have been discovered,but the pathogenesis and prognostic markers of CN-AML were not yet fully understood.In recent years, more and more studies have focused on the epigenetic regulation of AML.Non-coding RNA (such as lncRNA or miRNA) is an important part of the epigenetic regulation. Recent studies have shown that lncRNAs are closely related to tumor cell proliferation, invasion, metastasis, apoptosis and tumor angiogenesis. However, the specific regulatory mechanism of lncRNAs in CN-AML is still unclear. Since lncRNAs can regulate miRNAs abundance by binding and sequestering them which were knowed as “miRNA sponges”, lncRNAs can regulate the expression of target mRNAs.Thus, it has been shown that an efficient way to infer the potential function of lncRNAs is by studying their relationship with miRNAs and/or mRNAs, whose functions have been annotated.Therefore, this suggested that we can use the ceRNA network to explore the specific functional roles and prognostic significance of lncRNAs in CN-AML.
In this study, we conducted the difference analysis between CN-AML patients and normal controls to find differentially expressed RNAs by GEO data sets (GSE142699, GSE142698 and GSE103828) and document retrieval.Then the target lncRNAs and mRNAs of miRNAs were searched through the online databases. Afterwards,we took the intersection between the target RNAs with the above-mentioned differentially expressed RNAs to obtain DElncRNA–DEmiRNA and DEmiRNA–DEmRNA pairs, in which the expression of DEmiRNAs were negatively correlated with DElncRNA and DEmRNA.Finally ,a three-level network diagram of lncRNA–miRNA–mRNA was constructed.
We performed KEGG and GO analysis on the DEmRNAs in the lncRNA–miRNA–mRNA network.The pathway analysis showed that 29 pathways were significantly enriched(P-value<0.05).Pathways ‘PI3K-Akt signaling pathway’,‘MAPK signaling pathway’ and ‘Ras signaling pathway’, which have been shown to play important roles in AML,were invovled. Furthermore,108 GO terms were significantly enriched with P-value <0.05.These significant GO terms involved G1/S transition of mitotic cell cycle, cell cycle arrest and protein phosphorylation for the biological process group;protein binding, ATP binding and protein kinase activity for the molecular function group;and nucleus,nucleoplasm and cytosol for the cellular component group.
In order to find the key lncRNAs, which can be used as potential novel biomarkers for clinical diagnosis and treatment targets of CN-AML, the hub nodes and the number of relationship pairs were used. In this study, three lncRNAs (XIST、GABPB1-AS1、TUG1) were observed to be topological key nodes whose node degrees and the number of lncRNA–miRNA and miRNA–mRNA pairs were significantly higher compared to other lncRNAs. This indicated that these lncRNAs had profound implications for AML, which can be considered as key lncRNAs.
The XIST locus produces a 17–20 kb RNA that coats the X chromosome in cis, and plays an essential role in X chromosome inactivation (XCI).Much previous work has focused on the role of XIST in initiating X-inactivation in the early developing embryo. However,XIST has been reported to function as an oncogene or a tumor suppressor in different human malignancies, which is implicated in many aspects of carcinogenesis including tumor apoptosis, cell cycle, initiation, invasion, metastasis, stemness, autophagy and drug resistance.For example,XIST was highly expressed in breast cancer and was closely associated with a poor prognosis and the resistance to chemotherapy.Down-regulation of XIST has been reported to reduce chemoresistance in non-small cell lung cancer cells by inhibiting autophagy . In AML,XIST was found to be up-regulated in patients’bone marrow cells. In addition, silencing of XIST could repress AML bone marrow cell proliferation while enhancing cell apoptosis and adriamycin sensitivity.
In this study,lncRNA XIST was highly expressed in CN-AML and had the highest node degrees and the highest number of lncRNA–miRNA and miRNA–mRNA pairs in the ceRNA network among all the lncRNAs.This means XIST may play an important role in CN-AML which is consistent with previous findings mentioned above.The pathway analysis of key lncRNA XIST–miRNA–mRNA sub-network showed that 16 pathways were significantly enriched.Pathways‘PI3K-Akt signaling pathway’,‘FoxO signaling pathway’,‘p53 signaling pathway’ and ‘Ras signaling pathway’, which had been shown to play important roles in AML,were involved.
LncRNA GAbinding protein transcription factor subunit beta-1 antisense RNA 1 (GABPB1-AS1) is the antisense RNA of GABPB1 mRNA, which is located in the cytoplasm and has a total length of 4139nt.GABPB1-AS1 was identified for the first time in human-induced pluripotent stem cells (hiPSCs). The expression level of GABPB1-AS1 is increased in hiPSCs under the chemical stresses (cadmium, hydrogen peroxide, and cycloheximide).LncRNA GABPB1-AS1 aslo played a role in several cancers.But the results were contradictory.As a tumor suppressor gene, Qi et al found that GAPBPB1-AS1 inhibited the antioxidant ability of hepatocellular carcinoma cancer cells and cell proliferation by inhibiting the expression of GABPB1 and peroxiredoxin 5 (PRDX5).In addition,GABPB1-AS1 inhibited clear cell renal cell carcinoma growth and played a tumor suppressor role through an miR-1246/PCK1 axis.As a tumor activator gene, GABPB1-AS1 can bind to miR-519e-5p and destroied its tumor suppressive function in cervical cancer pathogenesis.Forthermore, findings suggested that the decrease in GABPB1-AS1 expression associated with decreased breast cancer risk.Alkhateeb et al also revealed that the aberrant expression of GABPB1-AS1 can be used as a potential prostate cancer biomarker.However, the role of GABPB1-AS1 in AML is still unclear.
In this study, for the first time, we came to a conclusion that lncRNA GABPB1-AS1 was highly expressed in CN-AML by both bioinformatics analysis and qRT-PCR verification in AML cell line(THP-1).The pathway analysis of key lncRNA GABPB1-AS1–miRNA–mRNA sub-network showed that 6 pathways were significantly enriched and primarily involved ‘PI3K-Akt signaling pathway’ and ‘Pathways in cancer’pathway terms.In addition,the survival analysis told us that patients with lower expression of GABPB1-AS1 had better prognosis.In conclusion,GABPB1-AS1 would like to be a potential prognostic marker and therapeutic target of AML.
LncRNA Taurine-Upregulated Gene1 (TUG1), located on chromosome 22q12, a critical oncogenic lncRNAs of human, has been proved to take part in hematological cancers. Interestingly, Some scholars demonstrated that TUG1 was highly expressed in tissues and cell lines of AML patients, and the high expression of TUG1 was also closely related to poor prognosis of AML.TUG1 induced cell proliferation, and restrained cell apoptosis in AML by targeting aurora kinase A.TUG1 facilitates the cell viability and metastasis by targeting miR-370-3p/MAPK1/ERK in AML.In addition,TUG1 silencing decreased the IC50 of adriamycin, and promotes adriamycin-induced apoptosis in AML cells by miR-34a/EZH2 axis,providing a potential therapeutic target for AML.
In our study,LncRNA TUG1 was highly expressed in CN-AML and had the higher topological parameters in the ceRNA network.This means TUG1 may play an important role in CN-AML which was consistent with previous findings mentioned above.The pathway analysis of key lncRNA TUG1–miRNA–mRNA sub-network showed that 16 pathways were enriched.Pathways ‘PI3K-Akt signaling pathway’,‘FoxO signaling pathway’,and ‘Ras signaling pathway’, which have been shown to play important roles in AML,were involved.
In summary, our research results constructed a lncRNA–miRNA–mRNA network associated with CN-AML, and provided novel lncRNAs(especially GABPB1-AS1) as potential diagnostic and prognostic biomarkers.The specific functional mechanism of these key lncRNAs in AML needs further experimental exploration.