MicroRNA‑425 upregulation indicates better prognosis in younger acute myeloid leukemia patients undergoing chemotherapy
- Authors:
- Published online on: April 4, 2019 https://doi.org/10.3892/ol.2019.10217
- Pages: 5793-5802
Abstract
Introduction
Acute myeloid leukemia (AML) is the most common type of acute leukemia affecting adults as a complex, dynamic disease (1). AML patients have a highly heterogeneous disease course, and the clinical outcomes are based on cytogenetic abnormalities and molecular genetic aberrations (2,3). For instance, NPM1 and biallelic CEBPA mutations are favorable prognostic markers for patients with cytogenetically normal AML (4). In addition, mutations of FLT3-ITD (5), DNMT3A (6), TP53 (7), RUNX1 (8) and MLL-PTD (9) consistently confer poor prognosis in AML patients. Recently, microRNAs (miRNAs) have been reported to serve an important role in the initiation, progression and prognosis of AML as epigenetic alterations (10).
miRNAs are evolutionary conserved, small (typically 18–25 nucleotides), non-coding RNAs that negatively regulate gene expression at the post-transcriptional level and play a crucial part in carcinogenesis. miRNAs not only function as tumor suppressors or oncogenes, but have also been implicated in cell migration and metastasis (11,12). In AML, changes in the expression of several miRNAs have been demonstrated to have functional relevance in leukemogenesis and supply prognostic information, complementing the evidence gained from cytogenetics, gene mutations and altered gene expression (10). For instance, it has been reported that overexpression of miR-191 and miR-199a contributes to an inferior outcome in AML (13), whereas increased expression of miR-181a and miR-212 predicts improved survival rates for AML patients (14,15).
The function of miR-425 has recently been reported in multiple human cancer types (16–19). Previous research has revealed that miR-425 functions as an oncogene or tumor suppressor in different cancer contexts. For instance, miR-425 upregulation promoted cell proliferation, migration and invasion in gastric cancer through a process involving CYLD (20), while it was also reported to inhibit cancer progression in melanoma via IGF-1 (21). However, the potential prognostic role and clinical implications of miR-425 in AML remain unclear.
The present study investigated whether the expression levels of miR-425 provide prognostic information for younger AML patients treated with chemotherapy, independently from a comprehensive panel of other established clinical and molecular predictors. The findings indicated that miR-425 may have future applications in guiding therapeutic interventions.
Patients and methods
Patients
The study included a total of 162 AML patients, whose information was retrieved from The Cancer Genome Atlas (TCGA) database (https://cancergenome.nih.gov/). The expression levels of miRNA-425, and the clinical and molecular information of the patients were publicly available from the TCGA website (22). Among the 162 patients, 90 were received chemotherapy-based consolidation as they were treated according to their respective situation, while the remaining 72 patients were treated with allogeneic hematopoietic stem cell transplantation (allo-HSCT). Gene and miRNA expression profiling was performed using HGU133 Plus 2.0 oligonucleotide microarrays (Affymetrix; Thermo Fisher Scientific, Inc., Waltham, MA, USA) and custom miRNA microarrays at diagnosis. Event-free survival (EFS) and overall survival (OS) were considered as endpoints, respectively. EFS was defined as the time from diagnosis until mortality, relapse or the absence of complete remission. OS was determined as the time from diagnosis to mortality, or the end of the follow-up.
Statistical analysis
The clinical and molecular characteristics of patients were summarized using descriptive statistics. Mann-Whitney U test was performed to compare differences in continuous variables, while Pearson's χ2 analysis was utilized to compare the differences in categorical variables. Kaplan-Meier survival curves and the log-rank test were used to compare patient survival. Cox proportional hazards model was used to evaluate miR-425 expression level as a predictor of clinical outcome in the context of other prognostic factors in univariate and multivariate analyses. These other prognostic factors included white blood cell (WBC) count (<20×109/l vs. ≥20×109/l), age (≥60 vs. <60 years), and FLT3-ITD, NPM1, DNMT3A, TP53, RUNX1, TET2, CEBPA, MLL-PTD and IDH1/2 mutations. Statistical analyses were performed with SPSS (version 22.0; IBM Corp, Armonk, NY, USA) and GraphPad Prism (version 7.0; GraphPad Software, Inc., La Jolla, CA, USA) software. The results in all analyses were considered as statistically significant when the two-tailed P-value was <0.05.
Results
Association of clinical and molecular characteristics with miR-425 expression levels in the chemotherapy and allo-HSCT groups
A total of 162 AML patients were divided into the chemotherapy and allo-HSCT groups. Next, each group was divided into two further subgroups based on the median expression level of miR-425. Patients with miR-425 expression levels that were higher or equal to the median value were included in the high miR-425 expression group, while the remaining patients were included in the low miR-425 expression. The median expression level was 3,709.321 (range, 481.232–19,682.91) in the chemotherapy group and 3,171.966 (range, 942.01–17,575.09) in the allo-HSCT group. The correlation of the miR-425 expression level with the clinical and molecular characteristics of patients is fully described in Table I.
In the chemotherapy group, patients with high miR-425 expression level exhibited a higher prevalence of inv(16)/CBFβ-MYH11 and low-risk disease, whereas the percentage of peripheral blood (PB) blasts, French-American-British (FAB) classification subtype M0, intermediate-risk disease, and DNMT3A, IDH1/2 and RUNX1 mutations were lower in these patients. There were no significant differences between the two expression groups in terms of age and gender distribution, WBCs, bone marrow (BM) blasts, FAB subtypes other than M0, karyotypes other than inv(16)/CBFβ-MYH11, low-risk disease, relapse rate, and FLT3-ITD, NPM1, CEBPA, MLL-PTD, NRAS/KRAS, TET2 and TP53 mutations.
In the allo-HSCT group, patients with high miR-425 expression level exhibited increased prevalence of inv(16)/CBFβ-MYH11 and MLL-PTD mutations, whereas the prevalence of trisomy 8 karyotype was lower in these patients. There were no significant differences between the two expression groups in terms of age and gender distribution, WBCs, BM blasts, PB blasts, FAB classification, risk stratification, frequent AML mutations (FLT3-ITD, NPM1, CEBPA, DNMT3A, IDH1/2, RUNX1, NRAS/KRAS, TET2 and TP53) and relapse rate.
Univariate and multivariate Cox analysis for prognosis in the chemotherapy and allo-HSCT groups
The effect of clinical and molecular characteristics on survival was next evaluated. The results of this analysis for the chemotherapy and allo-HSCT groups are summarized in Tables II and III.
 | Table III.Univariate and multivariate analyses for EFS and OS in the allogeneic hematopoietic stem cell transplantation group. |
In the chemotherapy group (Table II), univariate analysis revealed that high miR-425 expression was associated with significantly more favorable EFS (P=0.002) and OS (P=0.005), while TP53 mutations were associated with poor EFS (P=0.002) and OS (P=0.002). Since the age group was observed to be a significant predictive factor for EFS and OS (all P<0.001), younger and older subgroups were analyzed separately in multivariate analyses. In younger patients, only FLT3-ITD, NPM1, DNMT3A, TET2, IDH1/2 was included in multivariate analysis due to their relatively high mutation rate, which is more than 5% in younger patients. The results indicated that high miR-425 expression independently predicted a longer EFS and OS (both P=0.001). However, DNMT3A mutation in younger patients indicated a relatively shorter EFS (P=0.027) and OS (P=0.020), and IDH1/2 mutation also indicated shorter OS (P=0.044). In older patients, the miR-425 expression level was not associated with survival.
In the allo-HSCT group, univariate and multivariate analyses indicated that TP53 (P=0.015 and 0.008, respectively) and RUNX1 (P=0.038 and 0.021, respectively) mutations contributed to poor OS. In addition, MLL-PTD mutations had an adverse effect on EFS and OS in univariate analysis (both P<0.05), and remained significantly associated with shorter EFS in multivariate analysis (P=0.023). However, miR-425 had no effect on EFS and OS in univariate and multivariate analyses.
Subsequently, AML patients in both the chemotherapy and allo-HSCT groups were also analyzed as a whole using multivariate analysis, and the results are presented in Table IV. Mutations in DNMT3A and RUNX1 were observed to have an unfavorable effect on OS (P=0.031 and 0.012, respectively), while older age, chemotherapy and TP53 mutations contributed to poor EFS and OS (all P<0.05). No significant differences were identified in EFS and OS between the high and low miR-425 expression groups.
Prognostic value of miR-425 expression
Kaplan-Meier survival estimate in the chemotherapy group indicated a better prognosis for EFS (P<0.001) and OS (P=0.004) in patients with high expression of miR-425 as compared with that in patients exhibiting low miR-425 expression (Fig. 1A and B). Upon the division of AML patients undergoing chemotherapy into a younger and older age group, miR-425 was only associated with EFS and OS (both P=0.001) in younger patients (Fig. 1C and D), whereas no significant prognostic value was observed in older patients (Fig. 1E and F). In the allo-HSCT group, no significant differences were observed between patients with high versus low miR-425 expression (Fig. 1G and H).
Next, the entire cohort of patients was divided into two groups according to the expression levels of miR-425. Kaplan-Meier survival estimate demonstrated that no significant differences were observed between patients treated with allo-HSCT and chemotherapy in the high miR-425 expression group (Fig. 2A and B). By contrast, EFS and OS (both P<0.001) were longer in patients treated with allo-HSCT as compared with those treated with chemotherapy in the low miR-425 expression group (Fig. 2C and D).
Discussion
In the current study, higher miR-425 expression indicated better survival prospects for younger AML patients who received chemotherapy. By contrast, miR-425 expression exhibited no prognostic value in patients treated with allo-HSCT.
The data reported in the present study revealed that low-risk patients and the favorable cytogenetic alteration inv(16)/CBFβ-MYH11 appeared more frequently in the high miR-425 expression group, while unfavorable genetic mutations in RUNX1 were more often observed in the low expression group. This implies that miR-425 upregulation may serve the same role as inv(16)/CBFβ-MYH11 in predicting the prognosis for AML patients. Accordingly, downregulation of miR-425 may have similar prognostic features to RUNX1 mutation. Univariate analysis in the chemotherapy group indicated a putative favorable role of high miR-425 expression in AML patients. Furthermore, it was observed that the patient age had considerable implications on the therapeutic outcomes, and high miR-425 expression only indicated longer EFS and OS in younger AML patients that received chemotherapy. Kaplan-Meier survival curve analysis indicated the same results. By contrast, miR-425 expression levels were found to have no effect in the allo-HSCT group, suggesting that allo-HSCT overrides the prognostic ability of miR-425 expression.
Epigenetic modifiers, such as IDH1/2, TET2 and DNMT3A mutations, affect the expression of genes that are crucial to leukemogenesis, and as a consequence, they powerfully influence the prognosis of AML. In addition, IDH1, IDH2 and TET2 mutations are known to modulate DNA hydroxymethylation (23), while DNMT3A mutations are involved in DNA methylation, and increased risk of relapse or mortality in AML (24). It was demonstrated in the present study that the incidence of IDH1/2 and DNMT3A mutations was significantly higher in patients with low miR-425 expression, suggesting that miR-425 may also affect the prognosis through epigenetic regulation.
Allo-HSCT is one of the curative treatment options for patients with AML (25). In order to decide between transplant and non-transplant consolidation strategies, it is crucial to gain a clear idea of the outcome to be expected subsequent to allo-HSCT (26). The present study findings suggested that allo-HSCT may be more effective for AML patients expressing low miR-425 levels, whereas it may not be as effective for patients with high miR-425 expression, thus highlighting the potential utility of miR-425 in treatment selection.
There are certain limitations in the current study. Firstly, when patients treated with chemotherapy were analyzed by age subgroup, the sample size in each age group was small; in particular, there were only 29 patients in the younger subgroup. In addition, certain genes were required to be deleted from the multivariate analysis due to their low mutation rate, in order to ensure statistical efficiency. Finally, although the association between miR-425 expression levels and clinical outcomes was illustrated in this pilot study, further laboratory work is required to elucidate whether miR-425 functions as a tumor suppressor in AML and the underlying mechanisms involved. In our future work, in vitro and in vivo mouse experiments will be conducted to identify the target genes or pathways.
In conclusion, to the best of our knowledge, the present study analysis is the first to demonstrate that high miR-425 expression is an independent positive prognostic factor in younger AML patients undergoing chemotherapy. In addition, miR-425 upregulation may a factor for advising against allo-HSCT in AML patients.
Acknowledgements
Not applicable.
Funding
The present study was supported by grants from the National Natural Science Foundation of China (grant nos. 81500118 and 61501519), the China Postdoctoral Science Foundation Funded Project (grant no. 2016M600443) and the PLAGH Project of Medical Big Data (grant no. 2016MBD-025).
Availability of data and materials
The datasets analyzed during this study are available in The Cancer Genome Atlas database (https://cancergenome.nih.gov/).
Authors' contributions
LF and XK proposed and designed the study, and XK suggested analysis of the data based on age group of patients who underwent chemotherapy. JS screened and collected the data. XZ and XY were responsible for quality control of the data and performed the statistical analysis. GZ and JZ analyzed and interpreted the data, and JZ was a major contributor in writing the manuscript. KH performed the analysis, and generated the tables and figures. SY and JW interpreted the data, drafted the discussion, and revised and edited the entire manuscript. All authors read and approved the final manuscript.
Ethics approval and consent to participate
Not applicable.
Patient consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
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