Reversal of MYB-dependent suppression of MAFB expression overrides leukaemia phenotype in MLL-rearranged AML.

The transcription factor MYB plays a pivotal role in haematopoietic homoeostasis and its aberrant expression is involved in the genesis and maintenance of acute myeloid leukaemia (AML). We have previously demonstrated that not all AML subtypes display the same dependency on MYB expression and that such variability is dictated by the nature of the driver mutation. However, whether this difference in MYB dependency is a general trend in AML remains to be further elucidated. Here, we investigate the role of MYB in human leukaemia by performing siRNA-mediated knock-down in cell line models of AML with different driver lesions. We show that the characteristic reduction in proliferation and the concomitant induction of myeloid differentiation that is observed in MLL-rearranged and t(8;21) leukaemias upon MYB suppression is not seen in AML cells with a complex karyotype. Transcriptome analyses revealed that MYB ablation produces consensual increase of MAFB expression in MYB-dependent cells and, interestingly, the ectopic expression of MAFB could phenocopy the effect of MYB suppression. Accordingly, in silico stratification analyses of molecular data from AML patients revealed a reciprocal relationship between MYB and MAFB expression, highlighting a novel biological interconnection between these two factors in AML and supporting new rationales of MAFB targeting in MLL-rearranged leukaemias.

Ficoll-hypaque separation vs whole blood lysis: Comparison of efficiency and impact on minimal residual disease analysis.

INTRODUCTION: The high-throughput era remarkably changed molecular laboratory practice. Actually, the increasing number of processed samples requires to reduce the risk of operator biases, by automating or simplifying as much as possible both the analytical and the pre-analytical phases. Minimal residual disease (MRD) studies in hematology often require a simultaneous processing of many bone marrow and peripheral blood samples from patients enrolled in prospective, multicenter, clinical trials, monitored at several planned time points. METHODS: In this study, we demonstrate that red blood cell lysis (RBL) pre-analytical procedure can replace the time-consuming Ficoll stratification as cell recovering step. Here, we show a MRD comparison study using both total white blood cells and mononuclear cells recovered by the 2 procedures from 46 follicular lymphoma (FL), 15 multiple myeloma (MM), and 11 mantle cell lymphoma (MCL) patients enrolled in prospective clinical trials. RESULTS: The experiments were performed in the 4 laboratories of the Fondazione Italiana Linfomi (FIL) MRD Network and showed superimposable results, in terms of good correlation (R = 0.87) of the MRD data obtained by recovering blood cells by the 2 approaches. CONCLUSION: Based on these results, the FIL MRD Network suggests to optimize the pre-analytical phases introducing RBL approach for cell recovery in the clinical trials including MRD analysis.

Induction of sister-chromatid exchanges in human lymphocytes exposed in vitro and in vivo to therapeutic ultrasound.

The induction of sister-chromatid exchanges (SCEs), chromosomal aberrations and cell-cycle delay was determined in human lymphocytes after treatment in vitro and in vivo with therapeutic ultrasound (u.s.). In vitro treatments (1 W/cm2; 0.860 MHz; for 40-160 sec) were performed on unstimulated lymphocytes from 9 donors: a statistically significant, dose-dependent increase in SCE frequency was produced, whereas no induction of chromosomal aberrations nor alteration of the distribution of 1st, 2nd and 3rd division metaphases were observed. The same increase in the frequency of SCEs was detected by treating in vitro stimulated lymphocytes with u.s. The effects of in vivo exposure to u.s. were detected on lymphocytes from 10 patients before, during and after u.s. therapy (0.6-1.0 W/cm2; 0.860 MHz; from 8 to 20 applications lasting 5-6 min each). SCE frequency was statistically significantly increased in all patients at mid-therapy, without a further increase during the second half of therapeutic cycle, and was restored to pretreatment level 3 months after the end of u.s. therapy. No increase in chromosomal aberrations was noticed during and after u.s. therapy, whereas erratic delays of the cell cycle were observed, not clearly related to u.s. application or SCE levels. A linear relationship was found between SCE frequency and age in 21 healthy donors.