Copy Number Alterations are Associated with the Risk of Very Early Relapse in Pediatric B-lineage Acute Lymphoblastic Leukemia: A Nested Case-control MIGICCL Study.

BACKGROUND: Refining risk stratification to avoid very early relapses (VER) in Mexican patients with B-lineage acute lymphoblastic leukemia (B-ALL) could lead to better survival rates in our population. AIM OF THE STUDY: The purpose of this study was to investigate the association between the United Kingdom ALL (UKALL)-CNA classifier and VER risk in Mexican patients with childhood B-ALL. METHODS: A nested case-control study of 25 cases with VER and 38 frequency-matched controls without relapse was conducted within the MIGICCL study cohort. They were grouped into the categories of the UKALL-CNA risk classifier (good [reference], intermediate and poor), according to the results obtained by multiplex ligation dependent probe amplification. Overall and disease-free survival (DFS) were estimated using the Kaplan-Meier method. Univariate and multivariate Cox proportional hazards analyses were conducted. RESULTS: The CDKN2A/B genes were most frequently deleted in the group with relapse. According to UKALL-CNA classifier, 33 (52.4%) patients were classified as good, 21 (33.3%) intermediate and 9 (14.3%) poor-risk B-ALL. The intermediate and poor risk groups were associated with an increased risk of VER (HR = 4.94, 95% CI = 1.87-13.07 and HR = 7.42, 95% CI = 2.37-23.26, respectively) in comparison to the good-risk patients. After adjusting by NCI risk classification and chemotherapy scheme in a multivariate model, the risks remained significant. CONCLUSIONS: Our data support the clinical utility of profiling CNAs to potentially refine current risk stratification strategies of patients with B-ALL.

Identification of genomic copy number variations in lung benign metastasizing leiomyomatosis.

OBJECTIVES: Lung metastasizing leiomyomatosis (LML) is an infrequently diagnosed pathology developed after sexual maturation, commonly preceded by uterine myomas. Symptoms can include difficulties to breathe, cough, dyspnea and pain, because of mechanical obstruction exerted by expanding local growing leiomyomas. Lung leiomyomas are normally detected by imaging studies, but nowadays the precise diagnosis demands histological characterization of biopsies obtained from the affected tissues. The purpose of the present study was to determine the presence of genomic alterations in circulating cells of LML. METHODS: Immunohistochemical characterization of a lung biopsy extracted by thoracoscopy was performed. Pathologic proliferative smooth muscle cells were observed in a major lung metastasizing nodule, with a growing pattern similar to a uterine myoma. The presence of cellular linages different to smooth muscle cells was discarded by testing the presence of a battery of molecular markers. Also, a normal karyotype was determine by GTG-banding cytogenetic study, but a high density microarray analysis revealed six submicroscopic chromosomal regions displaying genomic abnormalities: microduplications were detected on chromosomes 4, 14, 17 and 22; and microdeletions on chromosomes 8 and 10. CONCLUSION: This study remarks the relevance of submicroscopic chromosomal analysis of unusual pathologic conditions such as Benign Metastasizing Leiomyomatosis. This propitiate a better understanding of the molecular basis on the development of the pathology, in order to reckon on minimally invasive diagnostic methods, and to design appropriate treatments.

miRNome landscape analysis reveals a 30 miRNA core in retinoblastoma.

BACKGROUND: miRNAs exert their effect through a negative regulatory mechanism silencing expression upon hybridizing to their target mRNA, and have a prominent position in the control of many cellular processes including carcinogenesis. Previous miRNA studies on retinoblastoma (Rb) have been limited to specific miRNAs reported in other tumors or to medium density arrays. Here we report expression analysis of the whole miRNome on 12 retinoblastoma tumor samples using a high throughput microarray platform including 2578 mature miRNAs. METHODS: Twelve retinoblastoma tumor samples were analyzed using an Affymetrix platform including 2578 mature miRNAs. We applied RMA analysis to normalize raw data, obtained categorical data from detection call values, and also used signal intensity derived expression data. We used Diana-Tools-microT-CDS to find miRNA targets and ChromDraw to map miRNAs in chromosomes. RESULTS: We discovered a core-cluster of 30 miRNAs that were highly expressed in all the cases and a cluster of 993 miRNAs that were uniformly absent in all cases. Another 1022 miRNA were variably present in the samples reflecting heterogeneity between tumors. We explored mRNA targets, pathways and biological processes affected by some of these miRNAs. We propose that the core-cluster of 30 miRs represent miRNA machinery common to all Rb, and affecting most pathways considered hallmarks of cancer. In this core, we identified miR-3613 as a potential and critical down regulatory hub, because it is highly expressed in all the samples and its potential mRNA targets include at least 36 tumor suppressor genes, including RB1. In the variably expressed miRNA, 36 were differentially expressed between males and females. Some of the potential pathways targeted by these 36 miRNAs were associated with hormonal production. CONCLUSION: These findings indicate that Rb tumor samples share a common miRNA expression profile regardless of tumor heterogeneity, and shed light on potential novel therapeutic targets such as mir-3613 This is the first work to delineate the miRNA landscape in retinoblastoma tumor samples using an unbiased approach.

Differentially Expressed Long Non-Coding RNAs Were Predicted to Be Involved in the Control of Signaling Pathways in Pediatric Astrocytoma.

Expression changes for long non-coding RNAs (lncRNAs) have been identified in adult glioblastoma multiforme (GBM) and in a mixture of adult and pediatric astrocytoma. Since adult and pediatric astrocytomas are molecularly different, the mixture of both could mask specific features in each. We determined the global expression patterns of lncRNAs and messenger RNA (mRNAs) in pediatric astrocytoma of different histological grades. Transcript expression changes were determined with an HTA 2.0 array. lncRNA interactions with microRNAs and mRNAs were predicted by using an algorithm and the LncTar tool, respectively. Interactomes were constructed with the HIPPIE database and visualized with the Cytoscape platform. The array showed expression changes in 156 and 207 lncRNAs in tumors (versus the control) and in pediatric GBM (versus low-grade astrocytoma), respectively. Predictions identified lncRNAs that have putative microRNA binding sites, which might suggest that they function as sponges in these tumors. Also, lncRNAs were shown to interact with many mRNAs, such as Pleckstrin homology-like domain, family A, member 1 (PHLDA1) and sulfatase 2 (SULF2). For example, qPCR found long intergenic non-coding RNA regulator of reprogramming (linc-RoR) expression levels upregulated in pediatric GBM when they were compared with control tissues or with low-grade tumors. Meanwhile, PHLDA1 and ELAV-like RNA binding protein 1 (ELAV1) showed expression changes in tumors relative to the control. Our data showed many lncRNAs with expression changes in pediatric astrocytoma, which might be involved in the regulation of different signaling pathways.

Copy Number Alterations Associated with Acute Lymphoblastic Leukemia in Mexican Children. A report from The Mexican Inter-Institutional Group for the identification of the causes of childhood leukemia.

B-cell precursor acute lymphocytic leukemia (B-ALL) represents a worldwide public health issue. Particularly, Mexico is one of the countries with the highest incidence of ALL in children. Between the multiple factors involved in ALL etiology, genetic alterations are clearly one of the most relevant features. In this work, a group of 24 B-ALL patients, all negative for the four most frequent gene fusions (ETV6-RUNX1, BCR-ABL1, TCF3-PBX1 and MLL-AF4), were included in a high-resolution microarray analysis in order to evaluate genomic copy-number alterations (CNAs). The results of this preliminary report showed a broad genomic heterogeneity among the studied samples; 58% of the patients were hyperdiploid and 33% displayed a chromosome 9p deletion of variable length affecting genes CDKN2A/B, two patients displayed genomic instability with a high number of focal CNAs, three patients presented unique duplications affecting 2q, 12p and 1q, respectively, and one patient displayed no copy number imbalances. The copy-number profile of 44 genes previously related to B-ALL was heterogeneous as well. Overall results highlight the need for a detailed description of the genetic alterations in ALL cancer cells in order to understand the molecular pathogenesis of the disease and to identify any prognostic markers with clinical significance.

Breast cancer cell line MDA-MB-231 miRNA profile expression after BIK interference: BIK involvement in autophagy.

B-cell lymphoma 2 (BCL2)-interacting killer (apoptosis inducing) (BIK) has been proposed as a tumor suppressor in diverse types of cancers. However, BIK's overexpression in breast cancer (BC) and in non-small lung cancer cells (NSCLCs), associated with a poor prognosis, suggests its participation in tumor progression. In this study, we evaluated the global expression pattern of microRNAs (miRNAs), messenger RNA (mRNA) expression changes in autophagy, and autophagic flux after BIK interference. BIK gene expression was silenced by small interfering RNA (siRNA) in BC cell MDA-MB-231, and BIK interference efficiency was tested by real-time PCR and by Western blotting. BIK expression levels decreased by 75 ± 18 % in the presence of 600 nM siRNA, resulting in the abolishment of BIK expression by 94 ± 30 %. BIK interference resulted in the overexpression of 17 miRNAs that, according to the DIANA-miRPath v3.0 database, are mainly implied in the control of cell signaling, gene expression, and autophagy. The autophagy array revealed downregulation of transcripts which participate in autophagy, and their interactome revealed a complex network, where hepatocyte growth factor-regulated tyrosine kinase substrate (HGS), α-synuclein (SNCA), unc-51-like autophagy activating kinase 1/2 (ULK1/2), and mitogen-activated protein kinase 3 (MAPK3) were shown to be signaling hubs. LC3-II expression-an autophagy marker-was increased by 169 ± 25 % after BIK interference, which indicates the involvement of BIK in autophagy. Altogether, our results indicate-for the first time-that BIK controls the expression of miRNAs, as well as the autophagic flux in MDA-MB-231 cells.

Evidence of Insulin Resistance and Other Metabolic Alterations in Boys with Duchenne or Becker Muscular Dystrophy.

Aim. Our aim was (1) to determine the frequency of insulin resistance (IR) in patients with Duchenne/Becker muscular dystrophy (DMD/BMD), (2) to identify deleted exons of DMD gene associated with obesity and IR, and (3) to explore some likely molecular mechanisms leading to IR. Materials and Methods. In 66 patients with DMD/BMD without corticosteroids treatment, IR, obesity, and body fat mass were evaluated. Molecules involved in glucose metabolism were analyzed in muscle biopsies. Results show that 18.3%, 22.7%, and 68% were underweight, overweight, or obese, and with high adiposity, respectively; 48.5% and 36.4% presented hyperinsulinemia and IR, respectively. Underweight patients (27.3%) exhibited hyperinsulinemia and IR. Carriers of deletions in exons 45 (OR = 9.32; 95% CI = 1.16-74.69) and 50 (OR = 8.73; 95% CI = 1.17-65.10) from DMD gene presented higher risk for IR than noncarriers. We observed a greater staining of cytoplasmic aggregates for GLUT4 in muscle biopsies than healthy muscle tissue. Conclusion. Obesity, hyperinsulinemia, and IR were observed in DMD/BMD patients and are independent of corticosteroids treatment. Carriers of deletion in exons 45 or 50 from DMD gene are at risk for developing IR. It is suggested that alteration in GLUT4 in muscle fibers from DMD patients could be involved in IR.

Analysis of polymorphisms in genes (AGT, MTHFR, GPIIIa, and GSTP1) associated with hypertension, thrombophilia and oxidative stress in Mestizo and Amerindian populations of México.

Several polymorphisms related to hypertension, thrombophilia, and oxidative stress has been associated with the development of cardiovascular disease. We analyzed the frequency of M235T angiotensinogen (AGT), A222V 5,10 methylenete-trahydrofolate reductase (MTHFR), L33P glycoprotein IIIa (GPIIIa), and I105V glutathione S-transferase P1 (GSTP1)} polymorphisms in 285 individuals belonging to Mexican-Mestizo and five Amerindian population from México, by real time PCR allelic discrimination. Allele and genotype frequencies were compared using chi square tests. All populations followed the Hardy Weinberg equilibrium for assay markers with the exception of the Triki, whose were in Hardy Weinberg dysequilibrium for the glutathione S-transferase P1 polymorphism. Interestingly, according to all the analyzed single nucleotide polymorphisms (SNPs), the Triki population was the most differentiated and homogeneous group of the six populations analyzed. A comparison of our data with those previously published for some Caucasian, Asian and Black populations showed quite significant differences. These differences were remarkable with all the Mexican populations having a lower frequency of the 105V allele of the glutathione S-transferase P1 and reduced occurrence of the 222A allele of the 5,10 methylenetetrahydrofolate reductase. Our results show the genetic diversity among different Mexican populations and with other racial groups.

Identification of duchenne muscular dystrophy female carriers by fluorescence in situ hybridization and RT-PCR.

Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disorder caused by mutations in the dystrophin DMD gene located at Xp21.1 region. Up to 65% of the patients present dystrophin gene deletions. Mothers of DMD patients have a two-thirds chance of carrying a dystrophin mutation. The female carrier will transmit the disease gene to half of her sons and half of her daughters. As the recurrence risk for the disease is extremely high, it is very important to detect carrier status among female relatives of the patients to bring an adequate genetic counseling. In this work, results from two methods to identify female carriers are presented. One method is a multicolor fluorescence in situ hybridization (FISH) assay, and the other is reverse transcriptase-polymerase chain reaction (RT-PCR). We showed that FISH is an efficient, sensitive method that brings confident results to detect DMD female carriers as compared to RT-PCR.