Selected miRNA levels are associated with IKZF1 microdeletions in pediatric acute lymphoblastic leukemia.

The clinical outcome of children with high-risk relapsed B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is poor. The present study assessed the utility and prognostic value of selected microRNA (miRNA/miR) in BCP-ALL. The changes in the expression levels of these miRNAs regarding known gene lesions affecting lymphoid development [early B-cell factor 1 (EBF1), ETS variant 6 (ETV6), IKAROS family zinc finger 1 (IKZF1), paired box 5 (PAX5), cyclin dependent kinase inhibitor (CDKN) 2A/CDKN2B, retinoblastoma 1 (RB1), pseudoautosomal region 1 (PAR1), B-cell translocation gene 1 protein (BTG1)] were analyzed. The following miRNAs were analyzed: miR-24, miR-31, miR-128, miR-542, and miR-708. The present study focused on patients with deletions of the IKAROS transcriptional factor gene IKZF1, which is currently considered to be an independent negative prognostic factor for ALL outcome. It was demonstrated that the expression level of miR-128 was significantly lower in patients with IKZF1 deletion compared with patients without IKZF1 deletion. Additionally, low expression of miR-542 was associated with CDKN2A/B and miR-31deletions, and low expression of miR-24 was associated with miR-31 deletion. Low expression of miR-31, miR-24, miR-708 and miR-128 was associated with PAX5 deletion, high expression of miR-24 and miR-542 was associated with PAR1 deletion and high expression of miR-708 was associated with ETV6 deletion. The expression of the selected miRNAs was not associated with deletions of BTG1, EBF1 and RB1. These data, by emphasizing the association of miRNAs expression level with microdeletions, may assist to elucidate ALL biology and contribute to future studies on the possible applications of the miRNA profile for diagnosis.

Genomic profiling of Acute lymphoblastic leukemia in ataxia telangiectasia patients reveals tight link between ATM mutations and chromothripsis.

Recent developments in sequencing technologies led to the discovery of a novel form of genomic instability, termed chromothripsis. This catastrophic genomic event, involved in tumorigenesis, is characterized by tens to hundreds of simultaneously acquired locally clustered rearrangements on one chromosome. We hypothesized that leukemias developing in individuals with Ataxia Telangiectasia, who are born with two mutated copies of the ATM gene, an essential guardian of genome stability, would show a higher prevalence of chromothripsis due to the associated defect in DNA double-strand break repair. Using whole-genome sequencing, fluorescence in situ hybridization and RNA sequencing, we characterized the genomic landscape of Acute Lymphoblastic Leukemia (ALL) arising in patients with Ataxia Telangiectasia. We detected a high frequency of chromothriptic events in these tumors, specifically on acrocentric chromosomes, as compared with tumors from individuals with other types of DNA repair syndromes (27 cases total, 10 with Ataxia Telangiectasia). Our data suggest that the genomic landscape of Ataxia Telangiectasia ALL is clearly distinct from that of sporadic ALL. Mechanistically, short telomeres and compromised DNA damage response in cells of Ataxia Telangiectasia patients may be linked with frequent chromothripsis. Furthermore, we show that ATM loss is associated with increased chromothripsis prevalence in additional tumor entities.

Germline activating TYK2 mutations in pediatric patients with two primary acute lymphoblastic leukemia occurrences.

The contribution of genetic predisposing factors to the development of pediatric acute lymphoblastic leukemia (ALL), the most frequently diagnosed cancer in childhood, has not been fully elucidated. Children presenting with multiple de novo leukemias are more likely to suffer from genetic predisposition. Here, we selected five of these patients and analyzed the mutational spectrum of normal and malignant tissues. In two patients, we identified germline mutations in TYK2, a member of the JAK tyrosine kinase family. These mutations were located in two adjacent codons of the pseudokinase domain (p.Pro760Leu and p.Gly761Val). In silico modeling revealed that both mutations affect the conformation of this autoregulatory domain. Consistent with this notion, both germline mutations promote TYK2 autophosphorylation and activate downstream STAT family members, which could be blocked with the JAK kinase inhibitor I. These data indicate that germline activating TYK2 mutations predispose to the development of ALL.

Preserved global histone H4 acetylation linked to ETV6-RUNX1 fusion and PAX5 deletions is associated with favorable outcome in pediatric B-cell progenitor acute lymphoblastic leukemia.

Epigenetic dysregulation is a hallmark of cancer executed by a number of complex processes the most important of which converge on DNA methylation and histone protein modifications. Epigenetic marks are potentially reversible and thus promising drug targets. In the setting of acute lymphoblastic leukemia (ALL) they have been associated with clinicopathological features including risk of relapse or molecular subgroups of the disease. Here, using immunocytochemistry of bone marrow smears from diagnosis, we studied global histone H4 acetylation, whose loss was previously linked to treatment failure in adults with ALL, in pediatric patients. We demonstrate that preserved global histone H4 acetylation is significantly associated with favorable outcome (RFS, EFS, OS) in children with B cell progenitor (BCP) ALL, recapitulating the findings from adult populations. Further, for the first time we demonstrate differential histone H4 acetylation in molecular subclasses of BCP-ALL including cases with ETV6-RUNX1 fusion gene or PAX5 deletion or deletions in genes linked to B cell development. We conclude global histone H4 acetylation is a prognostic marker and a potential therapeutic target in ALL.

Role of base-excision repair in the treatment of childhood acute lymphoblastic leukaemia with 6-mercaptopurine and high doses of methotrexate.

Methotrexate (MTX) and 6-mercaptopurine (6MP) are the most commonly used drugs in the therapy of childhood acute lymphoblastic leukaemia (ALL). The main genotoxic effect of MTX resulting from inhibition of thymidylate synthase is mis-incorporation of uracil into DNA, which is considered essential for the effectiveness of the Protocol M in ALL IC BFM 2002/EURO LB 2002 regimens. In this study, we investigated the level of basal and induced DNA damage as well as the effectiveness of DNA repair in lymphocytes of children with ALL at four time-points during therapy with MTX and 6MP. To assess DNA damage and the efficacy of DNA repair we used the modified alkaline comet assay with uracil DNA glycosylase (Udg) and endonuclease III (EndoIII). In addition, we examined the induction of apoptosis in the lymphocytes of the patients during treatment. Finally, we compared the activity of base-excision repair (BER), involved in removal of both uracil and oxidized bases from DNA in lymphocytes of children with ALL and lymphocytes of healthy children. BER efficiency was estimated in an in vitro assay with cellular extracts and plasmid substrates of heteroduplex DNA with an AP-site. Our results indicate that there is a significant decrease in the efficacy of DNA repair associated with an increased level of uracil in DNA and induction of apoptosis during therapy. Moreover, it was found that the BER capacity was decreased in the lymphocytes of ALL patients in contrast to that in lymphocytes of healthy children. Thus, we suggest that an impairment of the BER pathway may play a role in the pathogenesis and therapy of childhood ALL.