T-cell acute lymphoblastic leukemia (T-ALL) is characterized by genetic, immunophenotypic, and clinical heterogeneity. miRNAs are small non-coding RNAs involved in post-transcriptional silencing of gene expression, thus implicated in shaping the phenotype of cells, in health and disease. Aberrantly expressed miRNAs are implicated in haematological malignancies as either oncogenic or tumor suppressor miRNAs, collectively termed oncomiRs (miRNAs related to oncogenesis). Several miRNAs have been related to leukemia progression or treatment response. The profile of miRNA expression remains to be elucidated in pediatric T-ALL.

Next-generation sequencing (NGS) of the entire repertoire of expressed miRNAs (miRNA-seq) might aid the understanding of T-ALL biology and heterogeneity, providing novel candidate oncogenic/tumor suppressor miRNAs and miRNAs which might potentially serve as prognostic or classification markers in T-ALL.


We aimed to characterize miRNA expression profile in pediatric T-ALL with the focus on the identification of miRNA expression profiles reflecting T-ALL heterogeneity. We aimed to assess the potential application of NGS-based miRNA profiling for the classification of pediatric T-ALL. We also aimed to identify novel candidate oncomiRs implicated in T-ALL pathobiology (potential oncogenic and tumor suppressor miRNAs).


Study group consisted of 34 pediatric T-ALL patients and 5 age-related healthy bone marrow donors. Total RNA, including small RNA fraction, was isolated using miRCURY RNA Isolation Kit-Cell & Plant (Exiqon) from CD3+ cells selected immunomagnetically by negative selection with use of BD IMag Human T Lymphocyte Enrichment Set-DM (BD Biosciences). The libraries were prepared using the NEBNext Small RNA Library preparation kit (New England Biolabs). miRNA-seq was performed using NextSeq500 Illumina, with 10 mln reads/per sample, read length: 51 bp single-end; reference annotation: GRCh37, miRbase 20 (NGS Service Exiqon).


We showed that miRNA expression profile is discriminative between pediatric T-ALL and healthy bone marrow controls. We identified 23 miRNAs overexpressed in T-ALL vs. controls (including known and candidate novel oncogenic miRNAs in T-ALL). Among these are miRNAs with known mRNA target genes implicated in T-ALL pathogenesis, including: miR-20b-5p, miR-363-3p, miR-128-3p, miR-181b-5p, miR-181a-5p. We identified 38 underexpressed miRNAs (potential tumor suppressors). Among these are miRNAs with known T-ALL-implicated targets: miR-145-5p, miR-143-3p, miR-27a-5p, miR-24-3p, miR-10b-5p. We showed that miRNA expression profile is also discriminative between immunophenotypic subtypes of T-ALL (EGIL stages: II, III, IV), based on differential expression of 94 miRNAs.


NGS-based miRNA profiling provides insights into the heterogeneity of pediatric T-ALL at the miRNome level. Expression profiles of miRNAs differentially expressed in patients vs. controls and in T-ALL immunophenotypic subtypes, differing in their maturation stages (EGIL stages II, III, IV) hold potential for improved classification of T-ALL. Individual differentially expressed miRNAs are candidates for: functional analysis to assess their potential oncogenic/tumor suppressor role in T-ALL, and association analysis (survival and treatment response) to assess their potential as prognostic markers in this disease.

Research funded by National Science Centre, Poland grant: 2014/15/B/NZ2/03394 and National Centre of Research and Development (NCRD) grant STARTEGMED3/304586/5/NCBR/2017.


No relevant conflicts of interest to declare.

Author notes


Asterisk with author names denotes non-ASH members.