Clinical Study On The Methylation Pattern Of MiR-378 5’-Flanking Region In Myeloid Malignancies

Objective The present study was aimed to investigate the methylation status of mi R-378 5’-flanking region and to explore its clinical significance in patients with acute myeloid leukemia(AML), chronic myeloid leukemia(CML) and myelodysplastic syndrome(MDS).Methods Methylation status of mi R-378 5’-flanking region was investigated by real-time quantitative methylation-specific PCR(RQ-MSP) and bisulfite-sequencing PCR(BSP). The expression of mi R-378 was evaluated by real-time quantitative PCR(RQ-PCR).Results The hypomethylation of mi R-378 5’-flanking region was present in all three hematologic myeloid malignancies:(1) Hypomethylation of mi R-378 5’-flanking region was found in 54(41.5%) of 130 AML patients. The abnormal methylation of mi R-378 was not associated with the age, sex, hemoglobin level(HB), platelet counts(PLTs), white blood cell counts(WBCs), bone marrow blasts(BM blasts) of patients(P>0.05). There was no significant difference in hypomethylation incidence among FAB, WHO or karyotype subtypes(P>0.05), But the level of mi R-378 hypomethylation significantly increased in M2 subtype compared to other subtypes. Furthermore, patients with t(8;21) harbored the highest level of mi R-378 hypomethylation. However, there was no significant difference in complete remission and overall survival between patients with high and low mi R-378 methylation(P>0.05). The association of mi R-378 expression with methylation was not observed in AML patients, but mi R-378 expression in THP-1 line was increased while methylation status of mi R-378 5’-flanking region was decreased after 5-aza-d C treatment.(2) 19(36.5%) cases of 52 CML patients were observed with mi R-378 5’-flanking region hypomethylation. There was no differentiation between mi R-378-hypermethylation and mi R-378-hypomethylation in sex, age, HB, WBCs, PLTs, karyotype classifications and BCR-ABL transcript in CML patients(P>0.05). The frequencies of mi R-378 hypomethylation in patients in accelerated phase, in chronic phase, and in blast crisis were(2/7, 28.5%),(15/40, 38%) and(2/5,40%), respectively, and the difference was not statistically significant(P>0.05).(3) The rate of mi R-378 5’-flanking region hypomethylation was 21.1%(20/95) in MDS patients. However, the abnormal methylation of mi R-378 was not observed to be correlated with the age, WBCs, PLTs, HB of patients(P>0.05). The frequency of mi R-378 hypomethylation was significantly higher in male(15/55, 27.3%) patients than in female patients(4/40, 10.0%)(P = 0.040). There was no significant difference in hypomethylation incidence among FAB, WHO or IPSS subtypes(P>0.05). mi R-378 hypomethylated patients acquired significantly lower OS compared with those with hypermethylated in both whole MDS patients and MDS patients less than 60 years(P=0.036, P=0.027, respectively). Futhermore, multivariate analysis indicated that mi R-378 hypomethylation was an independent adverse prognostic factor in MDS patients <60 years.Conclusions(1) Hypomethylation of mi R-378 5’-flanking region is a common molecular event in patients with myeloid malignancies;(2) AML patients with t(8;21) harbored the highest level of mi R-378 hypomethylation;(3) Hypomethylation of mi R-378 5’-flanking region is an independent risk factor in young MDS patients(age<60y).