Global Levels Of Histone Modifications Research Articles

Chronic alcohol-induced long-lasting working memory deficits are associated with altered histone H3K9 dimethylation in the prefrontal cortex.

Epigenetic modifications have emerged as key contributors to the enduring behavioral, molecular and epigenetic neuroadaptations during withdrawal from chronic alcohol exposure. The present study investigated the long-term consequences of chronic alcohol exposure on spatial working memory (WM) and associated changes of transcriptionally repressive histone H3 lysine 9 dimethylation (H3K9me2) in the prefrontal cortex (PFC). Male C57BL/6 mice were allowed free access to either 12% (v/v) ethanol for 5 months followed by a 3-week abstinence period or water. Spatial WM was assessed through the spontaneous alternation T-maze test. Alcoholic and water mice received daily injections of GABAB agonist baclofen or saline during alcohol fading and early withdrawal. Global levels of histone modifications were determined by immunohistochemistry. Withdrawal mice displayed WM impairments along with reduced prefrontal H3K9me2 levels, compared to water-drinking mice. The withdrawal-induced decrease of H3K9me2 occurred concomitantly with increased level of permissive H3K9 acetylation (H3K9ac) in the PFC. Baclofen treatment rescued withdrawal-related WM deficits and fully restored prefrontal H3K9me2 and H3K9ac. Alcohol withdrawal induced brain region-specific changes of H3K9me2 and H3K9ac after testing, with significant decreases of both histone marks in the dorsal hippocampus and no changes in the amygdala and dorsal striatum. Furthermore, the magnitude of H3K9me2 in the PFC, but not the hippocampus, significantly and positively correlated with individual WM performances. No correlation was observed between H3K9ac and behavioral performance. Results also indicate that pre-testing intraperitoneal injection of UNC0642, a selective inhibitor of histone methyltransferase G9a responsible for H3K9me2, led to WM impairments in water-drinking and withdrawal-baclofen mice. Collectively, our results demonstrate that alcohol withdrawal induced brain-region specific alterations of H3K9me2 and H3K9ac, an effect that persisted for at least three weeks after cessation of chronic alcohol intake. The findings suggest a role for long-lasting decreased H3K9me2 specifically in the PFC in the persistent WM impairments related to alcohol withdrawal.

ChIPseqSpikeInFree: a ChIP-seq normalization approach to reveal global changes in histone modifications without spike-in.

The traditional reads per million normalization method is inappropriate for the evaluation of ChIP-seq data when treatments or mutations have global effects. Changes in global levels of histone modifications can be detected with exogenous reference spike-in controls. However, most ChIP-seq studies overlook the normalization that must be corrected with spike-in. A method that retrospectively renormalizes datasets without spike-in is lacking. ChIPseqSpikeInFree is a novel ChIP-seq normalization method to effectively determine scaling factors for samples across various conditions and treatments, which does not rely on exogenous spike-in chromatin or peak detection to reveal global changes in histone modification occupancy. Application of ChIPseqSpikeInFree on five datasets demonstrates that this in silico approach reveals a similar magnitude of global changes as the spike-in method does. St. Jude Cloud (https://pecan.stjude.cloud/permalink/spikefree) and St. Jude Github ( https://github.com/stjude/ChIPseqSpikeInFree). Supplementary data are available at Bioinformatics online.

Age, but Not Amyloidosis, Induced Changes in Global Levels of Histone Modifications in Susceptible and Disease-Resistant Neurons in Alzheimer's Disease Model Mice.

There is increasing interest in the role of epigenetic alterations in Alzheimer’s disease (AD). The epigenome of every cell type is distinct, yet data regarding epigenetic change in specific cell types in aging and AD is limited. We investigated histone tail modifications in neuronal subtypes in wild-type and APP/PS1 mice at 3 (pre-pathology), 6 (pathology-onset) and 12 (pathology-rich) months of age. In neurofilament (NF)-positive pyramidal neurons (vulnerable to AD pathology), and in calretinin-labeled interneurons (resistant to AD pathology) there were no global alterations in histone 3 lysine 4 trimethylation (H3K4me3), histone 3 lysine 27 acetylation (H3K27ac) or histone 3 lysine 27 trimethylation (H3K27me3) in APP/PS1 compared to wild-type mice at any age. Interestingly, age-related changes in the presence of H3K27ac and H3K27me3 were detected in NF-labeled pyramidal neurons and calretinin-positive interneurons, respectively. These data suggest that the global levels of histone modifications change with age, whilst amyloid plaque deposition and its sequelae do not result in global alterations of H3K4me3, H3K27ac and H3K27me3 in NF-positive pyramidal neurons or calretinin-labeled interneurons.

Global histone modification fingerprinting in human cells using epigenetic reverse phase protein array

The balance between acetylation and deacetylation of histone proteins plays a critical role in the regulation of genomic functions. Aberrations in global levels of histone modifications are linked to carcinogenesis and are currently the focus of intense scrutiny and translational research investments to develop new therapies, which can modify complex disease pathophysiology through epigenetic control. However, despite significant progress in our understanding of the molecular mechanisms of epigenetic machinery in various genomic contexts and cell types, the links between epigenetic modifications and cellular phenotypes are far from being clear. For example, enzymes controlling histone modifications utilize key cellular metabolites associated with intra- and extracellular feedback loops, adding a further layer of complexity to this process. Meanwhile, it has become increasingly evident that new assay technologies which provide robust and precise measurement of global histone modifications are required, for at least two pressing reasons: firstly, many approved drugs are known to influence histone modifications and new cancer therapies are increasingly being developed towards targeting histone deacetylases (HDACs) and other epigenetic readers and writers. Therefore, robust assays for fingerprinting the global effects of such drugs on preclinical cell, organoid and in vivo models is required; and secondly, robust histone-fingerprinting assays applicable to patient samples may afford the development of next-generation diagnostic and prognostic tools. In our study, we have used a panel of monoclonal antibodies to determine the relative changes in the global abundance of post-translational modifications on histones purified from cancer cell lines treated with HDAC inhibitors using a novel technique, called epigenetic reverse phase protein array. We observed a robust increase in acetylation levels within 2–24 h after inhibition of HDACs in different cancer cell lines. Moreover, when these cells were treated with N-acetylated amino acids in addition to HDACs, we detected a further increase in histone acetylation, demonstrating that these molecules could be utilized as donors of the acetyl moiety for protein acetylation. Consequently, this study not only offers a novel assay for diagnostics and drug screening but also warrants further research of the novel class of inexpensive, non-toxic natural compounds that could potentiate the effects of HDAC inhibitors and is therefore of interest for cancer therapeutics.

Spatial distribution of epigenetic modifications in Brachypodium distachyon embryos during seed maturation and germination.

Seed development involves a plethora of spatially and temporally synchronised genetic and epigenetic processes. Although it has been shown that epigenetic mechanisms, such as DNA methylation and chromatin remodelling, act on a large number of genes during seed development and germination, to date the global levels of histone modifications have not been studied in a tissue-specific manner in plant embryos. In this study we analysed the distribution of three epigenetic markers, i.e. H4K5ac, H3K4me2 and H3K4me1 in ‘matured’, ‘dry’ and ‘germinating’ embryos of a model grass, Brachypodium distachyon (Brachypodium). Our results indicate that the abundance of these modifications differs considerably in various organs and tissues of the three types of Brachypodium embryos. Embryos from matured seeds were characterised by the highest level of H4K5ac in RAM and epithelial cells of the scutellum, whereas this modification was not observed in the coleorhiza. In this type of embryos H3K4me2 was most evident in epithelial cells of the scutellum. In ‘dry’ embryos H4K5ac was highest in the coleorhiza but was not present in the nuclei of the scutellum. H3K4me1 was the most elevated in the coleoptile but absent from the coleorhiza, whereas H3K4me2 was the most prominent in leaf primordia and RAM. In embryos from germinating seeds H4K5ac was the most evident in the scutellum but not present in the coleoptile, similarly H3K4me1 was the highest in the scutellum and very low in the coleoptile, while the highest level of H3K4me2 was observed in the coleoptile and the lowest in the coleorhiza. The distinct patterns of epigenetic modifications that were observed may be involved in the switch of the gene expression profiles in specific organs of the developing embryo and may be linked with the physiological changes that accompany seed desiccation, imbibition and germination.

10th NTES Conference: Nickel and arsenic compounds alter the epigenome of peripheral blood mononuclear cells

The mechanisms that underlie metal carcinogenesis are the subject of intense investigation; however, data from in vitro and in vivo studies are starting to piece together a story that implicates epigenetics as a key player. Data from our lab has shown that nickel compounds inhibit dioxygenase enzymes by displacing iron in the active site. Arsenic is hypothesized to inhibit these enzymes by diminishing ascorbate levels – an important co-factor for dioxygenases. Inhibition of histone demethylase dioxygenases can increase histone methylation levels, which also may affect gene expression. Recently, our lab conducted a series of investigations in human subjects exposed to high levels of nickel or arsenic compounds. Global levels of histone modifications in peripheral blood mononuclear cells (PBMCs) from exposed subjects were compared to low environmentally exposed controls. Results showed that nickel increased H3K4me3 and decreased H3K9me2 globally. Arsenic increased H3K9me2 and decreased H3K9ac globally. Other histone modifications affected by arsenic were sex-dependent. Nickel affected the expression of 2756 genes in human PBMCs and many of the genes were involved in immune and carcinogenic pathways. This review will describe data from our lab that demonstrates for the first time that nickel and arsenic compounds affect global levels of histone modifications and gene expression in exposed human populations.

A Novel Microscopy-Based High-Throughput Screening Method to Identify Proteins That Regulate Global Histone Modification Levels

Posttranslational modifications of histones play an important role in the regulation of gene expression and chromatin structure in eukaryotes. The balance between chromatin factors depositing (writers) and removing (erasers) histone marks regulates the steady-state levels of chromatin modifications. Here we describe a novel microscopy-based screening method to identify proteins that regulate histone modification levels in a high-throughput fashion. We named our method CROSS, for Chromatin Regulation Ontology SiRNA Screening. CROSS is based on an siRNA library targeting the expression of 529 proteins involved in chromatin regulation. As a proof of principle, we used CROSS to identify chromatin factors involved in histone H3 methylation on either lysine-4 or lysine-27. Furthermore, we show that CROSS can be used to identify chromatin factors that affect growth in cancer cell lines. Taken together, CROSS is a powerful method to identify the writers and erasers of novel and known chromatin marks and facilitates the identification of drugs targeting epigenetic modifications.

The significance of epigenetic alterations in lung carcinogenesis

Lung cancer is recognized as a leading cause of cancer-related death worldwide and its frequency is still increasing. The prognosis in lung cancer is poor and limited by the difficulties of diagnosis at early stage of disease, when it is amenable to surgery treatment. Therefore, the advance in identification of lung cancer genetic and epigenetic markers with diagnostic and/or prognostic values becomes an important tool for future molecular oncology and personalized therapy. As in case of other tumors, aberrant epigenetic landscape has been documented also in lung cancer, both at early and late stage of carcinogenesis. Hypermethylation of specific genes, mainly tumor suppressor genes, as well as hypomethylation of oncogenes and retrotransposons, associated with histopathological subtypes of lung cancer, has been found. Epigenetic aberrations of histone proteins and, especially, the lower global levels of histone modifications have been associated with poorer clinical outcome in lung cancer. The recently discovered role of epigenetic modifications of microRNA expression in tumors has been also proven in lung carcinogenesis. The identified epigenetic events in lung cancer contribute to its specific epigenotype and correlated phenotypic features. So far, some of them have been suggested to be cancer biomarkers for early detection, disease monitoring, prognosis, and risk assessment. As epigenetic aberrations are reversible, their correction has emerged as a promising therapeutic target.

Abstract 1046: The histone methyl transferase MMSET/WHSC1 promotes an epithelial-mesenchymal transition and invasive properties of prostate cancer

Abstract Prostate cancer (PCa) remains the most common neoplasia diagnosed among men in the USA, and the second leading cause of cancer deaths. Recent evidence has indicated that epigenetic alterations contribute to the initiation and progression of PCa; however, the mechanisms by which chromatin modifiers influence this disease are still poorly understood. The histone methyl transferase MMSET (Multiple Myeloma Set Domain) is overexpressed in a number of different tumor types, including PCa. While the precise mechanism of MMSET action is still not well defined, it is believed to involve alteration of chromatin structure through dimethylation of lysine 36 on histone H3 (H3K36me2). Analysis of microarray databases indicates that MMSET expression rises in the progression from benign prostatic epithelium to localized and metastatic PCa. We found that PCa cell lines express significantly higher levels of MMSET compared to immortalized but non-transformed prostate cells. In t(4;14)+ myelomas, MMSET overexpression changes global levels of histone modifications, promoting H3K36me2, and repressing trimethylation of lysine 27 (H3K27me3). Similarly, H3K36me2 and H3K27me3 levels in PCa cell lines depended on the expression of MMSET, while depletion of MMSET in benign prostatic epithelial cells did not affect chromatin modifications. Knockdown of MMSET in DU145 and PC-3 tumor cells decreased soft agar colony formation and cell proliferation. Furthermore, inhibition of MMSET expression strikingly decreased cell migration and invasion, while overexpression of MMSET in immortalized non-transformed RWPE-1 cells promoted migratory and invasive properties. These findings suggest that high levels of MMSET play a pathogenic role in PCa progression. Overexpression of MMSET in RWPE-1 cells was accompanied by an epithelial to mesenchymal transition (EMT), characterized by a change in cell morphology and increased expression of mesenchymal markers vimentin and N-cadherin. Among a panel of EMT promoting genes analyzed, Twist1 expression was strongly regulated by MMSET both in the knockdown and the overexpression models. Chromatin immunoprecipitation analysis demonstrated that MMSET binds to the Twist1 locus, leading to an increase in H3K36me2 and a decrease in the repressive mark H3K27me3, suggesting a direct role of MMSET in regulation of Twist1. siRNA depletion of Twist1 in MMSET-overexpressing RWPE-1 cells blocked invasion, indicating that Twist1 was a critical target of MMSET responsible for the acquisition of EMT properties. Intriguingly, Twist1 expression was also elevated in a panel of myeloma cell lines overexpressing MMSET due to chromosomal translocation t(4;14). Collectively, these data suggest that MMSET has a critical role in cancer pathogenesis and progression through epigenetic regulation of metastasis-related genes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1046. doi:1538-7445.AM2012-1046

Global Levels of Histone Modifications in Peripheral Blood Mononuclear Cells of Subjects with Exposure to Nickel

Background: Occupational exposure to nickel (Ni) is associated with an increased risk for lung and nasal cancers. Ni compounds exhibit weak mutagenic activity, cause gene amplification, and disrupt cellular epigenetic homeostasis. However, the Ni-induced changes in global histone modification levels have only been tested in vitro.Objective: This study was conducted in a Chinese population to determine whether occupational exposure to Ni is associated with alterations of global histone modification levels and to evaluate the inter- and intraindividual variance of global histone modification levels.Method: Forty-five subjects with occupational exposure to Ni and 75 referents were recruited. Urinary Ni and global H3K4 trimethylation, H3K9 acetylation, and H3K9 dimethylation levels were measured in peripheral blood mononuclear cells (PBMCs) of subjects.Results: H3K4me3 was elevated in Ni-exposed subjects (0.25% ± 0.11%) compared with referents (0.15% ± 0.04%; p = 0.0004), and H3K9me2 was decreased (Ni-exposed subjects, 0.11% ± 0.05%; referents, 0.15% ± 0.04%; p = 0.003). H3K4me3 was positively (r = 0.4, p = 0.0008) and H3K9ac was negatively (r = 0.1, p = 0.01) associated with urinary Ni. Interindividual variances of H3K4me3, H3K9ac, and H3K9me2 were larger compared with intraindividual variance in both exposure test groups, resulting in reliability coefficients (an estimate of consistency of a set of measurements) of 0.60, 0.67, and 0.79 for H3K4me3, H3K9ac, and H3K9me2, respectively, for Ni-exposed subjects and of 0.75, 0.74, and 0.97, respectively, for referent subjects.Conclusion: The results of this study indicate that occupational exposure to Ni is associated with alterations of global histone modification levels and that measurements of global levels of histone modifications are relatively stable over time in human PBMCs.

Global histone H3 lysine 27 (H3K27) methylation levels and their prognostic relevance in renal cell carcinoma

To evaluate if histone H3 lysine 27 (H3K27) methylation plays a role in renal cell carcinoma (RCC) tissue and whether its expression is a predictor of cancer recurrence in RCC. A tissue microarray (TMA) with 193 RCC specimens (comprising 142 clear-cell, 31 papillary, 10 chromophobe and 10 sarcomatoid RCC), 10 oncocytoma tissue specimens and a TMA with 30 benign renal tissue samples were stained with antibodies against H3K27-monomethyl (H3K27me1), H3K27-dimethyl (H3K27me2) and H3K27-trimethyl (H3K27me3). Sections were scored according to staining intensity and the proportion of epithelial cells showing nuclear staining. H3K27 methylation levels were correlated with established clinical-pathological variables (tumour-node-metastasis [TNM] stage, Fuhrman grade) and progression-free/cancer-specific survival. H3K27me1/-me2/-me3 staining was significantly more intense in papillary RCC then in clear-cell RCC. H3K27me3 levels were higher in oncocytoma than in RCC. H3K27me1/-me2/-me3 methylation levels were inversely correlated with Fuhrman grading and pT-stage. Global H3K27me1/-me2/-me3 methylation levels were always higher in benign renal tissue than in RCC with tumour relapse (H3K27me1 P < 0.001, H3K27me2 P= 0.032, H3K27me3 P= 0.004). Progression-free survival was shorter in patients with lower levels of H3K27me1 and H3K27me3 in the univariate analysis. The newly created H3K27me score (combining the staining levels of the single modifications) was a significant and independent predictor of RCC progression-free survival. The present study on H3K27-methylation supports the hypothesis that global histone modifications are potential markers of cancer prognosis in RCC. One reason could be that decreased H3K27 indicates transcriptional activation and therefore predicts cancer activation.

Global histone H4K20 trimethylation predicts cancer-specific survival in patients with muscle-invasive bladder cancer

•To determine the role of global histone methylation as a prognostic parameter in patients with bladder cancer. •We used a tissue microarray with samples from patients with non-muscle-invasive bladder cancer (NMIBC; n= 161), muscle-invasive bladder cancer (MIBC, n= 127), normal urothelium (NU; n= 31) and bladder cancer metastases (METS; n= 31) to determine global histone methylation (me) levels at histone H3 lysine 4 (H3K4) and H4K20. •Global histone modification levels (H3K4me1, H3K4me3, H4K20me1, H4K20me2, and H4K20me3) were lower in bladder cancer samples than in NU tissue •Global levels of H3K4me1, H4K20me1, H4K20me2 and H4K20me3 were decreasing from NU over NMIBC and MIBC to METS. •H4K20me1 levels were increased in patients with NMIBC with advanced pTstage and less differentiated bladder cancer. •In patients with MIBC, pTstage was negatively correlated with H3K4me1, H4K20me1 and H4K20me2 levels. •H4K20me3 levels were significantly correlated in a univariate and multivariate model with bladder cancer-specific mortality after radical cystectomy in patients with MIBC. •Global histone methylation levels may help to identify patients with bladder cancer with poor prognosis after radical cystectomy.

Abstract 1996: Chronic arsenic exposure and changes in global levels of histone modifications in a human population in Bangladesh

Abstract Arsenic and arsenic compounds have been classified as group 1 human carcinogens by IARC. Naturally occurring inorganic arsenic, in particular, has been implicated in the development of human skin, lung, and bladder cancers and has also been associated with the development of liver and prostate cancers. However, despite being a formidable carcinogen, arsenic is a poor mutagen and it has been suggested that arsenic-induced carcinogenesis maybe governed by epigenetic mechanisms. We have detected differences in the levels of several histone epigenetic marks, such as H3K9acetylation, H3K9dimethylation and H3K27trimethylation in human peripheral mononuclear cell collected from individuals exposed to varying levels of inorganic arsenic in their drinking water. The changes in the global levels of these epigenetic marks were primarily measured using a novel ELISA method, while changes of arsenic-induced histone modifications in peripheral blood lymphocytes treated in vitro have also been examined via Mass Spectrometry and Western Blotting. Furthermore, we have also found a profound increase in the expression of histone genes after in vitro treatment of peripheral blood lymphocytes with sodium arsenite and will be examining this finding further. To the best of our knowledge, this study is the first in its class to measure arsenic-induced epigenetic modifications in humans and how they relate to changes in gene expression in a human population. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1996. doi:10.1158/1538-7445.AM2011-1996

Global histone acetylation levels: Prognostic relevance in patients with renal cell carcinoma

Epigenetic alterations play an important role in carcinogenesis. Recent studies have suggested that global histone modifications are predictors of cancer recurrence in various tumor entities. Global histone acetylation levels (histone H3 lysine 9 acetylation [H3K9Ac], histone H3 lysine 18 acetylation [H3K18Ac], total histone H3 acetylation [H3Ac] and total histone H4 acetylation [H4Ac]) were determined in patients with renal cell carcinoma (RCC) using immunohistochemistry in a tissue micro array with 193 RCC and 10 oncocytoma specimens. The histone acetylation pattern was not different among the diverse histological subtypes of RCC or oncocytoma samples. The H3Ac levels were inversely correlated with pT-stage (P = 0.005), distant metastasis (P = 0.036), Fuhrman grading (P = 0.001) and RCC progression (P = 0.029, hazard ratio 0.87). H4Ac deacetylation was correlated with pT-stage (P = 0.011) and grading (P = 0.029). H3K18Ac levels were an independent predictor of cancer-progression following surgery for localized RCC in the univariate (P = 0.001, hazard ratio 0.78) and multivariate (P = 0.005, hazard ratio 0.82) analysis. In conclusion, our study supports the concept of global histone modification levels as a universal cancer prognosis marker, and provides evidence for the use of histone deacetylases inhibitors as future drugs in the therapy of RCC.

Association of Global Levels of Histone Modifications with Recurrence-Free Survival in Stage IIB and III Esophageal Squamous Cell Carcinomas

This study was aimed at understanding the effects of histone modifications on recurrence-free survival (RFS) after esophagectomy in esophageal squamous cell carcinoma (ESCC). The acetylation of histone H3 lysine (H3K9Ac), histone H3 lysine 18 (H3K18Ac), and histone H4 lysine 12 (H4K12Ac), and the dimethylation of histone H3 lysine 9 (H3K9diMe) and histone H4 arginine 3 (H4R3diMe) were analyzed by immunohistochemistry in 237 ESCCs. The K-means clustering algorithm was used to identify unique patterns of histone modifications. At a median follow-up of 5.1 years, 109 (46%) of 237 patients had developed recurrence of disease. Mean global levels of H3K9Ac, H3K18Ac, H3K9diMe, H4K12Ac, and H4R3diMe were 81.5%, 65.1%, 80.3%, 45.9%, and 27.4%, respectively. In the analysis of individual histones, a 1% increase in the global level of H3K18Ac in pathologic stage III worsened RFS at 1.009 times [95% confidence interval (CI), 1.001-1.016; P = 0.03], after adjusting for age, sex, and operative method. Cluster analysis also showed significant effects of histone modifications on RFS. For stage IIB cancers, Cox proportional hazards analysis showed that RFS of cluster 1, with high global levels of H3K18Ac and H4R3diMe, was 2.79 times poorer (95% CI, 1.14-6.27; P = 0.008) than that of cluster 2, with low levels. RFS for stage III cancers was also poorer in cluster 1 than cluster 2 (adjusted hazard ratio, 2.42; 95% CI, 1.10-5.34; P = 0.02). In conclusion, the present study suggests that global levels of histone modifications in ESCC may be an independent prognostic factor of RFS.

Global levels of histone modifications predict prostate cancer recurrence

Epigenetic alterations such as DNA methylation and histone modifications play important roles in carcinogenesis. It was reported that global histone modification patterns are predictors of cancer recurrence in various tumor entities. Our study was performed to evaluate histone lysine (H(x)K(y)) and histone acetyl (H(x)Ac) modifications in prostate tissue. A tissue microarray with 113 prostate cancer (PCA), 23 non-malignant prostate tissues was stained with antibodies against H3K4 mono-(H3K4me1), di-(H3K4me2), tri-(H3K4me3) methylation, H3K9me1, H3K9me2, H3K9me3, H3 and H4 pan-acetylation (H3Ac, H4Ac). We also analyzed H3K4 methylation in patients with advanced PCA (hormone-refractory PCA-HRPC, n = 34; hormone-dependent PCA, n = 30). Sections were scored according the staining intensity and the proportion of epithelial cells showing nuclear staining. H3K4me1, H3K9me2, H3K9me3, H3Ac, and H4Ac were significantly reduced in PCA compared to non-malignant prostate tissue. H3Ac and H3K9me2 levels allowed discrimination of PCA and non-malignant prostate tissue highly specifically (>91%) and sensitively (>78%) as determined via ROC analyses (AUC >0.91). Histone lysine methylation and histone acetylation marks were correlated with clinical-pathological parameters (i.e., digital rectal examination, preoperative PSA, pT-stage, lymph node metastasis, Gleason score). In addition, H3K4me1 was a significant predictor of PSA recurrence following radical prostatectomy. H3K4me1, H3K4me2, and H3K4me3 levels were significantly increased in HRPC. Global histone modification levels may help to identify patients with adverse prognosis, and represent a target for the future therapy of PCA.

Global Levels of Histone Modifications Predict Prognosis in Different Cancers

Cancer cells exhibit alterations in histone modification patterns at individual genes and globally at the level of single nuclei in individual cells. We demonstrated previously that lower global/cellular levels of histone H3 lysine 4 dimethylation (H3K4me2) and H3K18 acetylation (ac) predict a higher risk of prostate cancer recurrence. Here we show that the cellular levels of both H3K4me2 and H3K18ac also predict clinical outcome in both lung and kidney cancer patients, with lower levels predicting significantly poorer survival probabilities in both cancer groups. We also show that lower cellular levels of H3K9me2, a modification associated with both gene activity and repression, is also prognostic of poorer outcome for individuals with either prostate or kidney cancers. The predictive power of these histone modifications was independent of tissue-specific clinicopathological variables, the proliferation marker Ki-67, or a p53 tumor suppressor mutation. Chromatin immunoprecipitation experiments indicated that the lower cellular levels of histone modifications in more aggressive cancer cell lines correlated with lower levels of modifications at DNA repetitive elements but not with gene promoters across the genome. Our results suggest that lower global levels of histone modifications are predictive of a more aggressive cancer phenotype, revealing a surprising commonality in prognostic epigenetic patterns of adenocarcinomas of different tissue origins.

Ventilatory function as a predictor of fatal stroke.

To investigate the relation between ventilatory function and subsequent mortality due to cerebrovascular disease. Prospective longitudinal study. A total of 18,403 male civil servants aged 40-64 years at entry examination for the Whitehall study. Mortality from cerebrovascular disease (ICD8 430-438) after 18 years of follow up. In all, 262 men with sinus rhythm at entry died due to stroke during the 18 years of follow up. Compared with men with a forced expiratory volume in one second of greater than or equal to 3.5 litres those with a value of less than 3.0 litres were almost twice as likely to die of cerebrovascular disease (rate ratio adjusted for age and systolic blood pressure = 1.88, 95% confidence interval 1.32 to 2.69). This increased risk occurred within each tertile of systolic blood pressure. Nested case-control analyses were used to control precisely for confounding effects of age, height, and smoking (by matching) and employment grade and physiological risk factors (by modelling). The effect of forced expiratory volume in one second was independent of age, height, smoking habits, employment grade, blood pressure, weight, cholesterol concentration, glucose tolerance, electrocardiographic abnormalities, history of chest pain, and history of intermittent claudication. Measurements of ventilatory function may assist clinical decisions about whether to treat mild hypertension. Impaired ventilatory function and stroke may share common causes.