Integrated Molecular Characterization of Testicular Germ Cell Tumors.

We studied 137 primary testicular germ cell tumors (TGCTs) using high-dimensional assays of genomic, epigenomic, transcriptomic, and proteomic features. These tumors exhibited high aneuploidy and a paucity of somatic mutations. Somatic mutation of only three genes achieved significance-KIT, KRAS, and NRAS-exclusively in samples with seminoma components. Integrated analyses identified distinct molecular patterns that characterized the major recognized histologic subtypes of TGCT: seminoma, embryonal carcinoma, yolk sac tumor, and teratoma. Striking differences in global DNA methylation and microRNA expression between histology subtypes highlight a likely role of epigenomic processes in determining histologic fates in TGCTs. We also identified a subset of pure seminomas defined by KIT mutations, increased immune infiltration, globally demethylated DNA, and decreased KRAS copy number. We report potential biomarkers for risk stratification, such as miRNA specifically expressed in teratoma, and others with molecular diagnostic potential, such as CpH (CpA/CpC/CpT) methylation identifying embryonal carcinomas.

Overexpression of mir-183 and mir-494 promotes proliferation and migration in human breast cancer cell lines.

Breast cancer (BC) is a leading cause of cancer-associated mortality in females worldwide. MicroRNAs (miRNAs or miRs), a type of non-coding RNA, have been reported to be important in the regulation of BC onset and progression. Several studies have implicated the role of miR-183 and miR-494 in different types of cancer. However, the biological functions of these miRNAs in BC remain largely unknown. In the present study, the expression of both miRNAs was assessed in the MDA-MB-231 and MDA-MB-468 BC cell lines. It was hypothesized that miR-183 and miR-494 serve an important role in regulating the expression of key genes associated with the metastatic phenotype of BC cells. To further understand their role, the expression of these miRNAs was restored in selected BC cell lines. Functional assays revealed that overexpression of miR-183 or miR-494 modulated the proliferation and migration of MDA-MB-231 and MDA-MB-468 cells in vitro. Additionally, retinoblastoma 1 (RB1) was identified to be a downstream target of both miRNAs by in silico analysis. Western blotting revealed that upregulation of miR-183 was associated with downregulation of RB1 protein in MDA-MB-231 cells. In conclusion, the present results support the hypothesis that miR-183 and miR-494 serve a pivotal role in BC metastasis, and that miR-183 may act as an oncogene by targeting RB1 protein in MDA-MB-231 cells.

Characteristics of the phenotypic abnormalities of bone marrow cells in childhood myelodysplastic syndromes and juvenile myelomonocytic leukemia.

BACKGROUND: Immunophenotyping of bone marrow (BM) hemopoietic precursors is useful for diagnosis of adult myelodysplastic syndrome (MDS), but data concerning pediatric patients are limited. We analyzed immunophenotypic features of BM cells at diagnosis of children who were referred to the Brazilian Pediatric Cooperative Group of Myelodysplastic Syndromes. METHODS: Diagnosis was based on clinical information, peripheral blood counts, BM cytology and cytogenetics. Patients with Down syndrome were excluded. Children with deficiency anemias or transitory neutropenias were used as controls (CTRLs). Immunophenotyping was performed on an eight-color antibody platform evaluating myelomonocytic maturation and progenitor cells. RESULTS: A total of 32 patients were examined: 6 refractory cytopenia of childhood [RCC]; 5 refractory anemia with excess of blasts [RAEB]; 8 refractory anemia with excess of blasts in transformation [RAEB-t]; 13 juvenile myelomonocytic leukemia [JMML] and 10 CTRLs. Median age was 66 months (RCC), 68 months (RAEB/RAEB-t), 29 months (JMML) and 70 months (CTRLs). Median number of phenotypic alterations was 4 (range 1-6) in RCC; 6 (range 2-11) in RAEB/RAEB-t and 6 (range 2-11) in JMML (P = 0.004). The percentage of CD34+ /CD117+ /CD13+ cells was 0.5% (range 0.1-2.8) in RCC; 4.2% (range 0.3-10.1) in RAEB/RAEB-t and 3.7 % (range 0.5-8.6) in JMML cases, compared with 0.7% (0.5-1.2) in CTRLs (P < 0.0005). Aberrancies in antigen expression of myeloid progenitors were seen in 63% of JMML and in 45% of RAEB/RAEB-t. CD34+ /CD19+ /CD10+ cells were decreased or absent in patients compared with age-matched controls. T lymphocytes were decreased in JMML. CONCLUSIONS: Phenotypic abnormalities were similar to those found in adult MDS. A decrease in B-cell precursors was observed especially in RAEB/RAEB-t. JMML and RAEB showed a similar pattern.

Sense-antisense pairs in mammals: functional and evolutionary considerations.

BACKGROUND: A significant number of genes in mammalian genomes are being found to have natural antisense transcripts (NATs). These sense-antisense (S-AS) pairs are believed to be involved in several cellular phenomena. RESULTS: Here, we generated a catalog of S-AS pairs occurring in the human and mouse genomes by analyzing different sources of expressed sequences available in the public domain plus 122 massively parallel signature sequencing (MPSS) libraries from a variety of human and mouse tissues. Using this dataset of almost 20,000 S-AS pairs in both genomes we investigated, in a computational and experimental way, several putative roles that have been assigned to NATs, including gene expression regulation. Furthermore, these global analyses allowed us to better dissect and propose new roles for NATs. Surprisingly, we found that a significant fraction of NATs are artifacts produced by genomic priming during cDNA library construction. CONCLUSION: We propose an evolutionary and functional model in which alternative polyadenylation and retroposition account for the origin of a significant number of functional S-AS pairs in mammalian genomes.

Aberrant methylation in pediatric myelodysplastic syndrome.

BACKGROUND: Aberrant methylation of gene promoter region is responsible for inappropriate gene silencing, and it has been associated to initiation and progression of cancer. Aberrant promoter methylation is frequently observed in adult patients with myelodysplastic syndrome (MDS), but in pediatric patients it has been poorly investigated. METHODS: We examined the promoter methylation status of 13 genes in bone marrow cells collected at diagnosis of 21 pediatric patients with MDS (subtype RAEB or RAEB-t). For this analysis, we performed sodium bisulfite treatment of genomic DNA, followed by methylation specific PCR (MSP). RESULTS: In pediatric MDS samples, we observed two genes frequently methylated: CALCA was methylated in 85.7% (18/21) of the analyzed samples and CDKN2B in 50% (6/12). CONCLUSIONS: Our findings indicate that CALCA and CDKN2B are frequently methylated in pediatric MDS. It suggests that aberrant methylation in pediatric MDS seems to be similar to adult MDS, thus pediatric patients could be also benefited with treatment using demethylating agents.

Hypermethylation of CpG island in the promoter region of CALCA in acute lymphoblastic leukemia with central nervous system (CNS) infiltration correlates with poorer prognosis.

Promoter hypermethylation occurs early in leukemogenesis and seems to be associated with poor prognosis in acute lymphoblastic leukemia (ALL). The methylation status of the promoter region of six genes was analyzed in 71 children with ALL using methylation specific PCR (MSP). Calcitonin (CALCA) and E-cadherin (CDH1) were the most frequently methylated genes in this group of patients. Considering the patients with central nervous system (CNS) infiltration, the estimated 2-year overall survival (OS) was 20% for those with methylation in CALCA promoter and 85% for those without (p=0.001). Our results suggest that the hypermethylation of CALCA promoter is a promising prognostic marker and may predict a higher risk for ALL patients with CNS infiltration.