Establishment of epigenetic markers to predict irradiation efficacy against oropharyngeal cancer

Irradiation, or chemoradiotherapy, is a curative treatment for oropharyngeal squamous cell carcinoma (OPSCC). Its invasiveness, however, can often negate its efficacy. Therefore, developing methods to predict which patients would benefit from irradiation is urgent. Promoter DNA hypermethylation was recently reported to correlate with favorable OPSCC prognosis. It is still unclear, however, whether there is an association between promoter DNA methylation and response to irradiation. In this study, we analyzed DNA methylation in the specimens from 40 OPSCC patients who had undergone irradiation, using the Infinium assay. Our results showed significant correlation between high levels of promoter DNA methylation and better response to treatment (P < 0.01). We used the 10 most differentially‐methylated genes between responders and non–responders to develop a panel of predictive markers for efficacy. Our panel had high sensitivity, specificity and accuracy (92%, 93% and 93%, respectively). We conducted pyrosequencing to quantitatively validate the methylation levels of 8 of the 10 marker genes (ROBO1, ULK4P3, MYOD1, LBX1, CACNA1A, IRX4, DPYSL3 and ELAVL2) obtained by Infinium. The validation by pyrosequencing showed that these 8 genes had a high prediction performance for the training set of 40 specimens and for a validation set of 35 OPSCC specimens, showing 96% sensitivity, 89% specificity and 94% accuracy. Methylation of these markers correlated significantly with better progression‐free and overall survival rates, regardless of human papillomavirus status. These results indicate that increased DNA methylation is associated with better responses to irradiation therapy and that DNA methylation can help establish efficacy prediction markers in OPSCC.     

DNA hypermethylation is associated with invasive phenotype of malignant melanoma

Tumor cell invasion is one of the key processes during cancer progression, leading to life-threatening metastatic lesions in melanoma. As methylation of cancer-related genes plays a fundamental role during tumorigenesis and may lead to cellular plasticity which promotes invasion, our aim was to identify novel epigenetic markers on selected invasive melanoma cells. Using Illumina BeadChip assays and Affymetrix Human Gene 1.0 microarrays, we explored the DNA methylation landscape of selected invasive melanoma cells and examined the impact of DNA methylation on gene expression patterns. Our data revealed predominantly hypermethylated genes in the invasive cells affecting the neural crest differentiation pathway and regulation of the actin cytoskeleton. Integrative analysis of the methylation and gene expression profiles resulted in a cohort of hypermethylated genes (IL12RB2, LYPD6B, CHL1, SLC9A3, BAALC, FAM213A, SORCS1, GPR158, FBN1 and ADORA2B) with decreased expression. On the other hand, hypermethylation in the gene body of the EGFR and RBP4 genes was positively correlated with overexpression of the genes. We identified several methylation changes that can have role during melanoma progression, including hypermethylation of the promoter regions of the ARHGAP22 and NAV2 genes that are commonly altered in locally invasive primary melanomas as well as during metastasis. Interestingly, the down-regulation of the methylcytosine dioxygenase TET2 gene, which regulates DNA methylation, was associated with hypermethylated promoter region of the gene. This can probably lead to the observed global hypermethylation pattern of invasive cells and might be one of the key changes during the development of malignant melanoma cells.     

Epigenetic alterations induced by ambient particulate matter in mouse macrophages

Respiratory mortality and morbidity has been associated with exposure to particulate matter (PM). Experimental evidence suggests involvement of cytotoxicity, oxidative stress, and inflammation in the development of PM‐associated pathological states; however, the exact mechanisms remain unclear. In the current study, we analyzed short‐term epigenetic response to PM₁₀ (particles with aerodynamic diameter less than 10 μm) exposure in mouse ascitic RAW264.7 macrophages (BALB/C Abelson murine leukemia virus‐induced tumor). Ambient PM₁₀ was collected using a high volume sampler in Little Rock, AR. Analysis revealed that PM₁₀ was composed mainly of Al and Fe, and the water soluble organic fraction was dominated by aliphatic and carbohydrate fragments and minor quantities of aromatic components. Exposure to PM₁₀ compromised the cellular epigenome at concentrations 10–200 µg/ml. Specifically, epigenetic alterations were evident as changes in the methylation and expression of repetitive element‐associated DNA and associated DNA methylation machinery. These results suggest that epigenetic alterations, in concert with cytotoxicity, oxidative stress, and inflammation, might contribute to the pathogenesis of PM‐associated respiratory diseases. Environ. Mol. Mutagen. 55:428–435, 2014. © 2014 Wiley Periodicals, Inc.     

Aging-like Spontaneous Epigenetic Silencing Facilitates Wnt Activation,Stemness, and BrafV600E-Induced Tumorigenesis

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Mitoepigenetics and drug addiction

Being the center of energy production in eukaryotic cells, mitochondria are also crucial for various cellular processes including intracellular Ca²⁺ signaling and generation of reactive oxygen species (ROS). Mitochondria contain their own circular DNA which encodes not only proteins, transfer RNA and ribosomal RNAs but also non-coding RNAs. The most recent line of evidence indicates the presence of 5-methylcytosine and 5-hydroxymethylcytosine in mitochondrial DNA (mtDNA); thus, the level of gene expression – in a way similar to nuclear DNA – can be regulated by direct epigenetic modifications. Up to now, very little data shows the possibility of epigenetic regulation of mtDNA. Mitochondria and mtDNA are particularly important in the nervous system and may participate in the initiation of drug addiction. In fact, some addictive drugs enhance ROS production and generate oxidative stress that in turn alters mitochondrial and nuclear gene expression. This review summarizes recent findings on mitochondrial function, mtDNA copy number and epigenetics in drug addiction.     

Transposable elements shape the human proteome landscape via formation of cis‐acting upstream open reading frames

Transposons are major drivers of mammalian genome evolution. To obtain new insights into the contribution of transposons to the regulation of protein translation, we here examined how transposons affected the genesis and function of upstream open reading frames (uORFs), which serve as cis‐acting elements to regulate translation from annotated ORFs (anORFs) located downstream of the uORFs in eukaryotic mRNAs. Among 39,786 human uORFs, 3,992 had ATG trinucleotides of a transposon origin, termed “transposon‐derived upstream ATGs” or TuATGs. Luciferase reporter assays suggested that many TuATGs modulate translation from anORFs. Comparisons with transposon consensus sequences revealed that most TuATGs were generated by nucleotide substitutions in non‐ATG trinucleotides of integrated transposons. Among these non‐ATG trinucleotides, GTG and ACG were converted into TuATGs more frequently, indicating a CpG methylation‐mediated process of TuATG formation. Interestingly, it is likely that this process accelerated human‐specific upstream ATG formation within transposon sequences in 5′ untranslated regions after divergence between human and nonhuman primates. Methylation‐mediated TuATG formation seems to be ongoing in the modern human population and could alter the expression of disease‐related proteins. This study shows that transposons have potentially been shaping the human proteome landscape via cis‐acting uORF creation.