Elena Serrano

Uniparental disomy may be associated with microsatellite instability in acute myeloid leukemia (AML) with a normal karyotype

The discovery of underlying genetic lesions helps to better understand the mechanisms of leukemogenesis and identify prognostic subgroups. Recent insights have allowed normal karyotype acute myeloid leukemia (AML) to be split into many molecular entities according to the genetic status of FLT3, NPM, CEBPA and MLL. Genome-wide single nucleotide polymorphism analysis was performed on 22 well-characterised AML patients with a normal karyotype. At the same time, microsatellite instability was investigated using a commonly used panel of polymorphic markers. Loss of heterozygosity (LOH) was found in 22.7% of cases without an associated copy number variation, suggesting that LOH represented an acquired partial uniparental disomy (aUPD) event. Three UPD+ cases harboured NPM mutations, associated with FLT3-ITD in two of them. An additional UPD patient had mutations both in CEBPA and in WT1. MSI was present at three loci in the three UPD+ cases (60%), whereas single locus MSI was present in three UPD- patients (17%). MSI involved the polymorphic PIG3 promoter in two UPD+ cases. It remains to be tested whether UPD and MSI association marks a common pathway of leukemogenesis.

Acute Myeloid Leukemia Subgroups Identified by Pathway-Restricted Gene Expression Signatures

Acute myeloid leukemia (AML) is a heterogeneous group of disorders characterized by abnormal proliferation of myeloid precursors and a maturation block. Underlying genetic lesions determine an altered expression program (transcriptosome) that can be studied in depth by massive technologies. Alternatively, we selected a pathway profiling strategy based on the current knowledge in order to stratify de novo AML patients and identify those cases which would potentially benefit from the use of new chemotherapeutic agents. One hundred and thirty-two RNA samples obtained from de novo adult AML patients were tested for FLT3, FLT3-LG, NDST1, HDAC2, ATRX, FOS, DNMT1, DNMT3A, DNMT3B, NBS1, RAD50, MRE11A, Meis1 and Meis2 expression using quantitative PCR (qPCR) assays. Clinical and biologic findings were correlated with expression results. Cluster analysis was also performed. FLT3 expression defined three subgroups of patients. The best outcome was found in the group with the lowest FLT3 expression. Intermediate levels of FLT3 were associated with the worst outcome. Patients with low levels of ATRX more frequently presented an adverse karyotype whereas cases with preserved ATRX levels showed an excellent outcome. In accordance with previous results, Meis1 downregulation is a useful surrogate marker indicating a good prognosis in AML patients. Simple qPCR platforms may help to identify different biologic subgroups in AML.

High expression of CEACAM6 and CEACAM8 mRNA in acute lymphoblastic leukemias

CEACAM family members are a set of widely expressed proteins involved in several biological functions, including cell adhesion, migration, signal transduction, and the regulation of gene expression. Abnormal overexpression and downregulation of some CEACAMs have been described in tumor cells. Monoclonal antibodies grouped in the CD66 cluster recognize CEACAM members. Ectopic CD66 expression is commonly detected in B-cell lineage acute lymphoblastic leukemia (ALL). To investigate the CEACAM messenger RNA (RNA) expression in leukemic blasts, we performed a quantitative polymerase chain reaction (RQ-PCR) analysis in purified RNA samples from a consecutive series of acute leukemias (135 patients). Most B-cell lineage ALL expressed CD66 (79.5%), whereas no single case of T-cell lineage ALL disclosed CD66 reactivity (0%). All the BCR-ABL+ ALL cases showed CD66 expression. CD66 was positive even in cases without CD10 expression (72.7%) and/or with MLL rearrangements. Despite the sharp contrast between T-ALL and B-ALL in CD66 reactivity, CEACAM patterns were comparable, and only minor differences for CEACAM1 and CEACAM8 were detected. All the leukemic samples showed overexpression of CEACAM6 and 8 when compared with normal granulocytes. These results were confirmed by dilutional experiments. The leukemic pattern paralleled the normal regenerating bone marrow with lower values for CEACAM1. In line with the results for CD66 reactivity, neoplastic cell lines had a uniform low expression of CEACAM family members. It remains to be investigated whether these CEACAM disturbances provide growth advantages to tumoral cells by inhibiting the anoikis process.

The FOXO1-miR27 tandem regulates myometrial invasion in endometrioid endometrial adenocarcinoma.

Micro-RNA (miRNA) signatures influence the prognosis of cancer, but little is known about their role in myometrial invasion in endometrioid endometrial adenocarcinoma (EEC). We studied miRNA expression signatures in noninvasive and invasive EEC focusing on the alteration of miR-27 and its main target, FOXO1 as well as their relationship with the clinicopathological parameters and other genetic alterations such as PIK3CA mutations. In 25 tumors and 5 normal endometria, unsupervised hierarchical clustering analysis showed that normal endometria and noninvasive EEC were grouped together and separately from invasive and advanced stage tumors. Of the 20 miRNAs differentially expressed in noninvasive (stage IA) and myoinvasive adenocarcinomas (stage IB and IC), miR27 was overexpressed in invasive adenocarcinomas, and its expression increased linearly according to stage. Results were validated by quantitative real-time reverse transcription polymerase chain reaction in an independent series of 44 EEC. By in situ hybridization, miR-27 expression was limited to the stroma. Using quantitative real-time reverse transcription polymerase chain reaction, the expression of proapoptotic transcription factor FOXO1 was down-regulated in invasive compared with noninvasive tumors. Furthermore, we found that the expression of active caspase 3 was higher in noninvasive than invasive EEC. When stratified by PIK3CA mutations, all invasive tumors down-regulated FOXO1, but only nonmutated adenocarcinomas showed miR-27 overexpression. In conclusion, we propose that the miR27-FOXO1 tandem inhibits apoptosis and represents an alternative pathway for tumor cell survival in PIK3CA-nonmutated EEC.

Epigenetic-based treatments emphasize the biologic differences of core-binding factor acute myeloid leukemias

Acute myeloid leukemia (AML) is a heterogeneous group of disorders characterized by an abnormal proliferation of the myeloid precursors and a maturation block. The most common chromosomal lesions in AML are the t(8;21) and inv(16). To better understand the leukemogenic mechanism of these fusion proteins, we performed gene expression studies in samples from (8;21), AML1 mutated and inv(16) patients, as well as from the Kasumi-1 cell line and a U937 cell line expressing the AML1-ETO fusion gene. To assess the influence of associated epigenetic lesions, we performed gene expression studies in Kasumi-1 cells and cells extracted from an Inv(16) patient, both treated with demethylating and HDAC inhibitor agents. Shared deregulated genes in the different types of core-binding factor leukemias were identified. We found a tight link between Inv(16) and mutant AML1 samples. Furthermore, some of the genes deregulated by the leukemogenic process reverted to their normal expression with demethylating and HDAC inhibitor treatment, highlighting the role of chromatin remodeling processes in AML.

K313dup is a recurrent CEBPA mutation in de novo acute myeloid leukemia (AML).

The CEBPA gene codes for a transcription factor that has a pivotal role in controlling proliferation and differentiation of myeloid progenitors. Acquired CEBPA mutations have been found in acute myeloid leukemias (AML) with a good prognosis, and most of these patients have a normal karyotype. In this paper, we report four cases that displayed the same K313dup in the CEBPA gene. All four had an AML-M1 with CD7 positivity and T-cell receptor gamma chain (TCR-γ) rearrangement. This mutation could represent nearly 10% of all CEBPA mutations described to date. K313dup disappeared in samples from patients in complete remission. In transfected cells, the K313dup mutant had reduced protein stability with respect to the wild-type protein. K313dup seems to be selected in leukemic cells, and its frequency in other AML series could be determined using the screening method reported in this paper.

Feasibility of the AML profiler (Skyline™ Array) for patient risk stratification in a multicentre trial: a preliminary comparison with the conventional approach

Deoxyribonucleic acid microarrays allow researchers to measure mRNA levels of thousands of genes in a single experiment and could be useful for diagnostic purposes in patients with acute myeloid leukaemia (AML). We assessed the feasibility of the AML profiler (Skyline™ Array) in genetic stratification of patients with de novo AML and compared the results with those obtained using the standard cytogenetic and molecular approach.

Normal Hypocretin-1 (Orexin A) Levels in Cerebrospinal Fluid in Patients with Idiopathic Intracranial Hypertension

AIM Low levels of hypocretin-1 (HC-1) have been associated with hypersomnia, obesity, depression, and chronic headaches. These conditions are frequently present in patients with idiopathic intracranial hypertension (IIH) and may be associated with abnormalities of the hypocretin system. The aim of this study was to determine HC-1 concentrations in cerebrospinal fluid (CSF) in a series of patients with IIH and to compare these concentrations with those in a control group with no neurological alterations. PATIENTS AND METHODS This prospective study included a cohort of 26 consecutive patients with IIH who were mostly women (25 vs. 1) with a mean age of 42.5 ± 13.2. CSF samples were obtained from a lumbar puncture performed between 08:00 and 10:00 a.m. HC-1 was determined by a competitive radioimmunoassay (RIA) using I(125) as the isotope. Samples of normal CSF were obtained during spinal anesthesia for urological, general or vascular surgery from 40 patients (10 women and 30 men with a mean age of 63.7 ± 14.8) with no previous neurological or psychiatric history, a normal neurological examination, and MMSE scores of ≥ 24. RESULTS No statistically significant differences were found between HC-1 levels in the CSF of patients with IIH (119.61 ± 21.63 pg/mL) and those of the control group (119.07 ± 20.30 pg/mL; p = 0.918). CONCLUSIONS HC-1 is not associated with the clinical symptoms present in patients with IIH.