Random DNA fragmentation allows detection of single-copy, single-exon alterations of copy number by oligonucleotide array CGH in clinical FFPE samples

Genomic technologies, such as array comparative genomic hybridization (aCGH), increasingly offer definitive gene dosage profiles in clinical samples. Historically, copy number profiling was limited to large fresh-frozen tumors where intact DNA could be readily extracted. Genomic analyses of pre-neoplastic tumors and diagnostic biopsies are often limited to DNA processed by formalin-fixation and paraffin-embedding (FFPE). We present specialized protocols for DNA extraction and processing from FFPE tissues utilizing DNase processing to generate randomly fragmented DNA. The protocols are applied to FFPE clinical samples of varied tumor types, from multiple institutions and of varied block age. Direct comparative analyses with regression coefficient were calculated on split-sample (portion fresh/portion FFPE) of colorectal tumor samples. We show equal detection of a homozygous loss of SMAD4 at the exon-level in the SW480 cell line and gene-specific alterations in the split tumor samples. aCGH application to a set of archival FFPE samples of skin squamous cell carcinomas detected a novel hemizygous deletion in INPP5A on 10q26.3. Finally we present data on derivative of log ratio, a particular sensitive detector of measurement variance, for 216 sequential hybridizations to assess protocol reliability over a wide range of FFPE samples

Pathway Implications of Aberrant Global Methylation in Adrenocortical Cancer.

Adrenocortical carcinomas (ACC) are a rare tumor type with a poor five-year survival rate and limited treatment options.Understanding of the molecular pathogenesis of this disease has been aided by genomic analyses highlighting alterations in TP53, WNT, and IGF signaling pathways. Further elucidation is needed to reveal therapeutically actionable targets in ACC.In this study, global DNA methylation levels were assessed by the Infinium HumanMethylation450 BeadChip Array on 18 ACC tumors and 6 normal adrenal tissues. A new, non-linear correlation approach, the discretization method, assessed the relationship between DNA methylation/gene expression across ACC tumors.This correlation analysis revealed epigenetic regulation of genes known to modulate TP53, WNT, and IGF signaling, as well as silencing of the tumor suppressor MARCKS, previously unreported in ACC.DNA methylation may regulate genes known to play a role in ACC pathogenesis as well as known tumor suppressors

cDNA microarrays detect activation of a myogenic transcription program by the PAX3-FKHR fusion oncogene

U radu će biti prikazani načini upotrebe šumske biomase (drvne tvari) u kogeneracijskim postrojenjima gdje se drvna tvar koristi kako energent njezinim sagorijevanjem ili rasplinjavanjem (pirolizom). Isto tako će biti shematski prikazana kogeneracijska postrojenja sa pojašnjenjem svih faza rada. Na kraju rada će biti prikazani primjeri kogeneracijskih postrojenja koja su u funkciji na području RH sa njihovim osnovnim karakteristikama

Box plots display examples of genes significantly hyper or hypomethylated in ACC.

<p><i>EPHX3</i> and <i>MEIS</i> genes are significantly hypermethylated in ACC compared with <i>TMEM132D</i> and <i>ADCY2</i> which are significantly hypomethylated.</p

Unsupervised clustering analysis of normal and ACC samples using DML.

<p>Clustering analysis revealed separation of ACC and normal samples. Samples are on the horizontal axis: normal samples are shown with a yellow bar and ACC samples are shown with a blue bar.</p

Significantly regulated genes in the WNT pathway.

<p>Significantly regulated genes in the WNT pathway.</p

Methylation/expression correlations according to discretization method.

<p>For each gene, the upper heatmap represents the log2 methylation values for 14 ACC samples each normalized to the average of 6 normal adrenal samples. Log2 methylation ratios >0 represent hypermethylation and <0 represent hypomethylation. The lower heat map shows expression of z-transformed expression levels, where a value 0 indicates no expression change compared to average expression level of 14 ACC samples. Three genes with negative correlations were selected for visualization: <i>CCDC8</i> (TP53 pathway), <i>TBX3</i> (WNT pathway), and <i>PAX8</i> (other known cancer gene, involved in invasion and migration). Samples with higher methylation (peach-maroon) had lower expression (green). Samples with lower methylation (blue) had higher expression (red). Only the correlated methylation loci and expression probes are shown, and samples are organized by their discretization classification of M or U for each gene.</p

Frequency of DML by chromosome after normalization to chromosome length.

<p>Frequency of DML by chromosome after normalization to chromosome length.</p