The Role Of Non-Coding RNAs In The Classification And Prognosis Of Myelodysplastic Syndromes

Objective To investigate the expression difference of non-coding RNAs(ncRNAs)long non-coding RNAs(lncRNAs)and microRNAs(miRNAs)in different subtypes of myelodysplastic syndromes(MDS),and to verify the correlation of key miRNAs with clinical classification and prognosis.To analysis the potential regulatory mechanisms of MDS-related molecules and to predict their values in clinic.Method Based on the World Health Organization(WHO)classification of MDS(2016),140 bone marrow samples and clinical data(including routine blood test results,morphology and chromosome examination results of bone marrow,gene test results,etc.)of MDS patients were collected from the Institute of Hematology and Blood Diseases Hospital,Chinese Academy of Medical Sciences and Peking Union Medical College for retrospective study.Total RNA was isolated from bone marrow mononuclear cells(BMCs)and RNA sequencing was performed in 5 MDS with multilineage dysplasia(MDS-MLD)samples and 5 MDS with excess blasts(MDS-EB)samples.The differentially expressed miRNAs and lncRNAs between MDS-MLD and MDS-EB groups were screened out by bioinformation analysis,two of candidate miRNAs were selected for the further verification.The expression levels of miRNAs in all samples were detected by Quantitative Real-time PCR(qRT-PCR).According to the International Prognostic Scoring System(IPSS),WHO Classification-Based Prognostic Scoring System(WPSS)and Revised IPSS(IPSS-R),patients were divided into different groups by the risk grade,and the expression levels of key miRNAs in different risk groups and different subtypes were compared,the correlation between the expression of two miRNAs and some prognostic factors(white blood cell,hemoglobin,platelet,Neutrophils,bone marrow blasts and gene mutation)were analyzed.Using TargetScan to predict the potential target genes of miRNA and then do the Gene Ontology(GO)analysis,and the interaction among lncRNAs,miRNAs and mRNAs was also predicted.Result 1.37 miRNAs(20 up and 17 down)and 231 lncRNAs(150 up and 81 down)that significantly differential expression between MDS-MLD and MDS-EB groups were identified,and two key candidate molecules miR-141 and miR-181b were selected 2.Relative expression levels of miR-141 and miR-181b were significantly different in different classification(MDS-MLD and MDS-EB)of MDS(P<0.05),the expression of miR-18 1b in MDS-EB was significantly higher than that in MDS-MLD,on the contrary,the expression of miR-141 in MDS-EB was significantly lower than that in MDS-MLD.3.The expression of miR-181b was positively correlated with the scores of three MDS prognostic scoring systems(IPSS,WPSS and IPSS-R;r=0.448,0.345,0.283,P<0.01),and expression level of miR-181b was increased with the risk grade,the differences in groups were statistically significant(P<0.05);There was no correlation between the expression of miR-141 and the scores of three MDS prognosis scoring systems,and there was no significant difference of miR-141 expression level in different risk grades(P>0.05).4.miR-18 1b expression was positively correlated with the bone marrow blasts(r=0.506,P<0.01),and there was no significant relationship with Hb,PLT,ANC,and abnormal karyotype(P>0.05);miR-141 expression was negatively correlated with the ANC(r=-0.222,P=0.013),and there was no significant relationship with bone marrow blasts,Hb,PLT,and abnormal karyotype(P>0.05).5.A total of 1363 potential target genes of miR-18 1b were predicted by TargetScan,functions of these target genes were mainly enriched in transcription regulation,RNA metabolism regulation and so on.Among them,22 target genes were related to the hematological malignancies,including RUNX1,ASXL2,NRAS,ATM and KRAS which have been previously confirmed to be related to MDS.6.4 interrelated lncRNAs and 14 potential target mRNAs of miR-18 1b in MDS-EB were predicted.Conclusion There are significantly different expression of miRNAs and lncRNAs between MDS-MLD and MDS-EB,suggesting that the pathogenesis of the two subtypes may be distinct,and both miRNAs and lncRNAs have regulatory functions.The expression levels of miR-141 and miR-181b in MDS-MLD and MDS-EB are significantly different,revealing that they could be important regulatory factors of MDS and applicable for the classification.Expression level of miR-181b has a significant correlation with risks and some prognostic factors,which means it has a potential to become a molecular prognostic factor of MDS.