Gene expression analysis of microRNA molecules is becoming increasingly important. In this study we assess the use of the mean expression value of all expressed microRNAs in a given sample as a normalization factor for microRNA real-time quantitative PCR data and compare its performance to the currently adopted approach. We demonstrate that the mean expression value outperforms the current normalization strategy in terms of better reduction of technical variation and more accurate appreciation of biological changes.

T-cell acute lymphoblastic leukemia (T-ALL) is a high-risk subtype of acute lymphoblastic leukemia (ALL) with gradually improved survival through introduction of intensified chemotherapy. However, therapy-resistant or refractory T-ALL remains a major clinical challenge. Here, we evaluated B-cell lymphoma (BCL)-2 inhibition by the BH3 mimetic ABT-199 as a new therapeutic strategy in human T-ALL. The T-ALL cell line LOUCY, which shows a transcriptional program related to immature T-ALL, exhibited high in vitro and in vivo sensitivity for ABT-199 in correspondence with high levels of BCL-2. In addition, ABT-199 showed synergistic therapeutic effects with different chemotherapeutic agents including doxorubicin, l-asparaginase, and dexamethasone. Furthermore, in vitro analysis of primary patient samples indicated that some immature, TLX3- or HOXA-positive primary T-ALLs are highly sensitive to BCL-2 inhibition, whereas TAL1 driven tumors mostly showed poor ABT-199 responses. Because BCL-2 shows high expression in early T-cell precursors and gradually decreases during normal T-cell differentiation, differences in ABT-199 sensitivity could partially be mediated by distinct stages of differentiation arrest between different molecular genetic subtypes of human T-ALL. In conclusion, our study highlights BCL-2 as an attractive molecular target in specific subtypes of human T-ALL that could be exploited by ABT-199.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive form of leukemia that is mainly diagnosed in children and shows a skewed gender distribution toward males. In this study, we report somatic loss-of-function mutations in the X-linked histone H3K27me3 demethylase ubiquitously transcribed X (UTX) chromosome, in human T-ALL. Interestingly, UTX mutations were exclusively present in male T-ALL patients and allelic expression analysis revealed that UTX escapes X-inactivation in female T-ALL lymphoblasts and normal T cells. Notably, we demonstrate in vitro and in vivo that the H3K27me3 demethylase UTX functions as a bona fide tumor suppressor in T-ALL. Moreover, T-ALL driven by UTX inactivation exhibits collateral sensitivity to pharmacologic H3K27me3 inhibition. All together, our results show how a gender-specific and therapeutically relevant defect in balancing H3K27 methylation contributes to T-cell leukemogenesis. Key Points The H3K27me3 demethylase UTX is recurrently mutated in male T-ALL and escapes X-inactivation in female T-ALL blasts and normal T cells. The loss of Utx contributes to T-ALL formation in vivo and UTX inactivation confers sensitivity to H3K27me3 inhibition.

L-Asparaginase (L-ASNase) is an enzyme that hydrolyses the amino acid asparagine into aspartic acid and ammonia. Systemic administration of bacterial L-ASNase is successfully used to lower the bioavailability of this non-essential amino acid and to eradicate rapidly proliferating cancer cells with a high demand for exogenous asparagine. Currently, it is a cornerstone drug in the treatment of the most common pediatric cancer, acute lymphoblastic leukemia (ALL). Since these lymphoblasts lack the expression of asparagine synthetase (ASNS), these cells depend on the uptake of extracellular asparagine for survival. Interestingly, recent reports have illustrated that L-ASNase may also have clinical potential for the treatment of other aggressive subtypes of hematological or solid cancers. However, immunogenic and other severe adverse side effects limit optimal clinical use and often lead to treatment discontinuation. The design of optimized and novel L-ASNase formulations provides opportunities to overcome these limitations. In addition, identification of multiple L-ASNase resistance mechanisms, including ASNS promoter reactivation and desensitization, has fueled research into promising novel drug combinations to overcome chemoresistance. In this review, we discuss recent insights into L-ASNase adverse effects, resistance both in hematological and solid tumors, and how novel L-ASNase variants and drug combinations can expand its clinical applicability.