DLX4 hypermethylation is a prognostically adverse indicator in de novo acute myeloid leukemia

Abstract

Hypermethylation of distal-less homeobox 4 (DLX4) has been increasingly identified in several cancers. Our study was aimed to determine the role of DLX4 methylation in regulating DLX4 expression and further analyze its clinical significance in de novo acute myeloid leukemia (AML) patients. DLX4 methylation level was detected by real-time quantitative methylation-specific PCR and bisulfite sequencing PCR. Treatment with 5-aza-2'-deoxycytidine (5-aza-dC) was used for demethylation studies. Clinical significance of DLX4 methylation was obtained by the comparison between the patients with and without DLX4 methylation. DLX4 was significantly methylated in AML patients compared with controls (P < 0.001). DLX4 methylation was negatively associated with DLX7 (the shorter DLX4 isoform) (R = -0.202, P = 0.021) but not BP1 (the longer DLX4 isoform) (R = -0.049, P = 0.582) expression in AML patients. DLX7 and BP1 messenger RNA (mRNA) were significantly increased after 5-aza-dC treatment in leukemic cell lines THP1 and Kasumi-1. DLX4 methylated patients showed significantly higher frequency of U2AF1 mutation compared with DLX4 unmethylated patients (P = 0.043). Both all AML and non-M3 patients with DLX4 methylation presented significantly lower complete remission rate than those with DLX4 unmethylation (P = 0.001 and <0.001, respectively). DLX4 methylated cases had significantly shorter overall survival than DLX4 unmethylated cases among both all AML (P = 0.003), non-M3 AML (P = 0.001), and cytogenetically normal AML (P = 0.032). Multivariate analysis confirmed that DLX4 methylation was independent risk factor in both all AML and non-M3 patients. Our study indicates that DLX4 hypermethylation is negatively associated with DLX7 expression and predicts poor clinical outcome in de novo AML patients.

Epigenetic dysregulation of ID4 predicts disease progression and treatment outcome in myeloid malignancies

“…Recently, RNA splicing factors gene U2AF1 mutation could cause splicing alterations in biological pathways previously implicated in myeloid malignancies, including the DNA damage response and epigenetic regulation usually in DNA methylation through DNMT3B pathway [48]. Moreover, our previous study also found that GPX3 hypermethylation was correlated with CEBPA wild-type in AML, while DLX4 hypermethylation was associated with U2AF1 mutation [20,32]. However, the underlying mechanism of the relation between ID4 methylation CEBPA and/or U2AF1 mutation remains unknown.…”

“…The treatment for MDS patients with lower IPSS scores (Low/Int‐1) was symptomatic and supportive treatment with/without thalidomide, whereas patients with higher IPSS scores (Int‐2/High) received chemotherapy included aclacinomycin, cytarabine, granulocyte colony stimulating factor together with symptomatic and supportive treatment. Patients with AML received chemotherapy including induction therapy and subsequent consolidation treatment . For non‐M3 patients, induction therapy was one or two courses of daunorubicin combined with cytarabine.…”

“…Genomic DNA isolation and modification were performed as reported previously . RQ‐MSP was applied to detect the level of ID4 methylation using AceQ qPCR SYBR Green Master Mix (Vazyme Biotech Co., Piscataway, NJ, USA) with primers reported previously .…”

“…at 72°C. BSP products cloning sequencing was performed as described . Five independent clones from each specimen were sequenced (BGI Tech Solutions Co., Shanghai, China).…”

Methylation‐independent ITGA2 overexpression is associated with poor prognosis in de novo acute myeloid leukemia

“…MSP was applied to detect ITGA2 methylation using Takara Taq™ Hot Start Version (Tokyo, Japan) with primers presented in Table 2. were conducted as reported previously (Zhou et al, 2016). Six independent clones from each specimen were sequenced (BGI Tech Solutions Co., Shanghai, China).…”