Identification of a neocentromere in a rearranged Y chromosome with no detectable DYZ3 centromeric sequence

An 18-year-old woman was evaluated because of primary amenorrhea and hypogonadism. Chromosome analysis from peripheral blood lymphocytes revealed a nonmosaic 46,X,+mar constitution. The marker was shown to be a rearranged Y chromosome consisting of an inverted duplication of the long arm: rea(Y)(qter-q11113326326

Myelodysplastic Syndrome (MDS) Displays Profound and Functionally Significant Epigenetic Deregulation Compared to Acute Myeloid Leukemia (AML) and Normal Bone Marrow Cells

Abstract MDS are a clinically heterogeneous group of clonal disorders, which share a high frequency of progression to secondary AML (MDS-AML). In contrast to de novo AML, MDS and MDS-AML are uniformly resistant to conventional chemotherapy. Among the few active drugs in MDS are the nucleoside analog DNA methyltransferase inhibitors (MTIs) 5-azacytidine and decitabine. Since tumor suppressor genes such as CDKN2B can be silenced by DNA methylation in MDS, it is believed that reversal of DNA methylation by MTIs might contribute to their anti-tumor effects. However, it is not clear whether methylation-dependent silencing of specific genes are predictive of response or even correlate with response to MTIs. Like MDS, AML also presents epigenetic silencing of CDKN2B and other genes; yet appear to not be as sensitive to MTIs. Given the particular sensitivity of MDS to MTIs and its resistance to standard AML chemotherapy, we hypothesized that MDS is a biologically distinct disease from AML due largely to extensive epigenetic deregulation, which is missed by single locus studies. In order to test this we studied DNA methylation levels at 24,000 gene promoters in 13 MDS pts. and 16 de novo normal karyotype AML cases, and compared and contrasted these to CD34+ bone marrow cells from 8 healthy donors. For this we used the HELP (HpaII tiny fragment Enrichment by Ligation-mediated PCR) assay, a robust method for detection of whole-genome DNA methylation, and using MassArray quantitative methylation for single locus validation. Remarkably, MDS was found to have a far greater number of hypermethylated genes than AML or normal CD34+ cells, while AML and CD34+ cells had similar number of methylated genes (MDS vs. AML: 6303 vs. 4177 promoters, p=0.026; MDS vs. normal CD34+ cells: 6303 vs. 4296 promoters, p=0.056). Using a moderated T test, an aberrant methylation signature of 736 genes (p<0.0000001) was identified in MDS vs. normal CD34+ cells, reflecting extensive epigenetic deregulation in this disease, including p16, CEBPZ, MSH2, CHES1, AKT1, Caspase2, BMP3, DAP and MYOD1. A comparison between MDS and de novo AML identified 498 genes (p<0.00005) differentially methylated, with a clear predominance of hypermethylated promoters in MDS vs. de novo AML. These genes included RUNX2 and 3, GFI1, DAPK2, MDM2, TGFA, CEBPZ and SHARP. Finally, a comparison between normal CD34+ cells and de novo AML demonstrated an aberrant pattern of methylation in 341 genes (p<0.00001), including CXCL1 and 5, PPARD, Caspase 2 HOXA4 and HOXA10, GFI1 and the MLL translocation partner Septin11. 7 of the 13 MDS cases were also examined for gene expression using the Affymetrix Hgu133plus2 array. A significant proportion of the genes that had been found to be methylated were also found to be underexpressed, including p16, DAP, BMP3, HOXA2 and HOXB8. Taken together, our data show that MDS is a unique and distinct biological entity than de novo AML featuring profound and functionally significant genome wide epigenetic deregulation. While the de novo AML methylation profile was clearly different from normal CD34+ cells, it was not as severely altered as MDS. These data also suggest that MTIs are most likely uniquely active in MDS due to their DNA methyltransferase activity

Early-onset breast cancer in a woman with a germline mobile element insertion resulting in BRCA2 disruption : a case report

Mobile element insertions (MEIs) contribute to genomic diversity, but they can be responsible for human disease in some cases. Initial clinical testing (BRCA1, BRCA2 and PALB2) in a 40-year-old female with unilateral breast cancer did not detect any pathogenic variants. Subsequent reanalysis for MEIs detected a novel likely pathogenic insertion of the retrotransposon element (RE) c.7894_7895insSVA in BRCA2. This case highlights the importance of bioinformatic pipeline optimization for the detection of MEIs in genes associated with hereditary cancer, as early detection can significantly impact clinical management.Published versio

A Pathologic Link between Wilms Tumor Suppressor Gene, WT1, and IFI1612

The Wilms tumor gene (WT1) is mutated or deleted in patients with heredofamilial syndromes associated with the development of Wilms tumors, but is infrequently mutated in sporadic Wilms tumors. By comparing the microarray profiles of syndromic versus sporadic Wilms tumors and WT1-inducible Saos-2 osteosarcoma cells, we identified interferon-inducible protein 16 (IFI16), a transcriptional modulator, as a differentially expressed gene and a candidate WT1 target gene. WT1 induction in Saos-2 osteosarcoma cells led to strong induction of IFI16 expression and its promoter activity was responsive to the WT1 protein. Immunohistochemical analysis showed that IFI16 and WT1 colocalized in WT1-replete Wilms tumors, but not in normal human midgestation fetal kidneys, suggesting that the ability of WT1 to regulate IFI16 in tumors represented an aberrant pathologic relationship. In addition, endogenous IFI16 and WT1 interacted in vivo in two Wilms tumor cell lines. Furthermore, IFI16 augmented the transcriptional activity of WT1 on both synthetic and physiological promoters. Strikingly, short hairpin RNA (shRNA)-mediated knockdown of either IFI16 or WT1 led to decreased growth of Wilms tumor cells. These data suggest that IFI16 and WT1, in certain cellular context including sporadic Wilms tumors, may support cell survival

Growth Suppression by Acute Promyelocytic Leukemia-Associated Protein PLZF Is Mediated by Repression of c-myc Expression

The transcriptional repressor PLZF was identified by its translocation with retinoic acid receptor alpha in t(11;17) acute promyelocytic leukemia (APL). Ectopic expression of PLZF leads to cell cycle arrest and growth suppression, while disruption of normal PLZF function is implicated in the development of APL. To clarify the function of PLZF in cell growth and survival, we used an inducible PLZF cell line in a microarray analysis to identify the target genes repressed by PLZF. One prominent gene identified was c-myc. The array analysis demonstrated that repression of c-myc by PLZF led to a reduction in c-myc-activated transcripts and an increase in c-myc-repressed transcripts. Regulation of c-myc by PLZF was shown to be both direct and reversible. An interaction between PLZF and the c-myc promoter could be detected both in vitro and in vivo. PLZF repressed the wild-type c-myc promoter in a reporter assay, dependent on the integrity of the binding site identified in vitro. PLZF binding in vivo was coincident with a decrease in RNA polymerase occupation of the c-myc promoter, indicating that repression occurred via a reduction in the initiation of transcription. Finally, expression of c-myc reversed the cell cycle arrest induced by PLZF. These data suggest that PLZF expression maintains a cell in a quiescent state by repressing c-myc expression and preventing cell cycle progression. Loss of this repression through the translocation that occurs in t(11;17) would have serious consequences for cell growth control