Serum Let-7b and AKT3 predict cardiovascular death in type 2 diabetes: insights from miRNA profiling and target prediction
H E Evirgen, A S Hocaoglu, S Ahmadova, M Postula, S De Rosa, M Di Martino, C Eyileten, Z WicikAbstract
Background
Type 2 diabetes (T2D) is associated with increased cardiovascular (CV) morbidity and mortality. MicroRNAs (miRNAs) potential biomarkers for CV complications in T2D. This study investigated the prognostic value of circulating miRNAs and their mRNA targets for long-term CV mortality in high-risk T2D patients.
Methods
We studied 262 T2D patients with ≥2 CV risk factors (age 30–80), among whom 23 (9%) died of CV causes over ~6 years. Global miRNA and gene expression profiling (microarrays on 8 decedents vs 8 survivors) was conducted to discover differentially expressed miRNAs and genes. Bioinformatic analyses included miRNA target prediction (multiMiR) and application of the OneR machine-learning algorithm to identify top miRNAs clustering by the number of cardiovascular-related gene targets. Gene enrichment (EnrichR) and interaction network analyses (Cytoscape) were performed to reveal affected pathways and hub genes. Key miRNAs (e.g. let-7b-5p, miR-16-5p) and a target gene (AKT3) were validated by quantitative PCR in the full cohort. Prognostic performance was assessed using ROC curves, Kaplan–Meier survival analysis, and Cox regression modeling.
Results
Several circulating miRNAs were dysregulated in patients who died vs. survivors, including higher baseline levels of let-7b-5p, miR-16-5p, and AKT3 mRNA in decedents. Let-7b-5p and AKT3 emerged as the strongest predictors of CV mortality. Individually, low let-7b-5p and low AKT3 levels (below optimized cut-offs) were associated with ~7–8-fold higher hazard of CV death in Cox analysis (p<0.001). A combined biomarker panel (let-7b-5p + AKT3) improved risk prediction, yielding an area under the ROC curve of ~0.81 (74% sensitivity/74% specificity), outperforming either marker alone. Network analysis identified central gene targets of the dysregulated miRNAs (PTEN, CCND1, MDM2), with AKT3 linked to the AGE–RAGE signaling pathway in diabetes. Enrichment analysis indicated that the target genes of these miRNAs are involved in pathways related to metabolism, inflammation, and gene regulation (e.g. PTEN/PI3K–AKT signaling), providing insight into the mechanisms connecting the identified biomarkers to CV outcomes.
Conclusions
Circulating miRNAs and their gene targets can serve as novel prognostic biomarkers for CV mortality in patients with T2D at high risk. In particular, the combination of let-7b-5p and AKT3 expression stratified patients by mortality risk with robust predictive performance. These results suggest that integrating miRNA and mRNA expression profiling improves risk assessment and reveals underlying pathways (such as insulin/AKT signaling and inflammatory pathways) that contribute to adverse CV outcomes. Clinically, such biomarkers could inform more personalized management of high-risk T2D patients, although external validation is warranted before routine use.