Molecular, clinical and therapeutic determinants of outcome in NPM1 mutated AML.

Although NPM1-mutated acute myeloid leukemia (AML) carries a generally favorable prognosis, many patients still relapse and die. Previous studies identified several molecular and clinical features associated with poor outcome, however only FLT3-ITD mutation and adverse karyotype are currently used for risk stratification due to inconsistent results and uncertainty around how other factors should influence treatment, particularly given the strong prognostic impact of post-induction measurable residual disease (MRD). Here we analyzed a large group of patients with NPM1mut AML enrolled in prospective trials (NCRI AML17 and AML19, n=1357) to delineate the impact of baseline molecular and clinical features, post induction MRD status and treatment intensity on outcome. FLT3-ITD (HR 1.28, 95%CI 1.01-1.63), DNMT3A (HR 1.65, 95%CI 1.32-2.05), WT1 (HR 1.74, 95%CI 1272-2.38) and non-ABD NPM1 mutations (HR 1.64, 95%CI 1.22-2.21) were independently associated with poorer overall survival (OS). These factors were also strongly associated with MRD positivity. For patients achieving MRD negativity, these mutations (except FLT3-ITD) were associated with an increased cumulative incidence of relapse (CIR) and poorer OS. However, apart from the few patients with adverse cytogenetics, we could not identify any group of MRD negative patients with a CIR >40% or with benefit from allograft in first remission. Intensified chemotherapy with the FLAG-Ida regimen was associated with improved outcomes in all subgroups, with greater benefits observed in the highest risk molecular subgroups.

Study protocol: Minimum effective low dose: anti-human thymocyte globulin (MELD-ATG): phase II, dose ranging, efficacy study of antithymocyte globulin (ATG) within 6 weeks of diagnosis of type 1 diabetes.

INTRODUCTION: Type 1 diabetes (T1D) is a chronic autoimmune disease, characterised by progressive destruction of the insulin-producing ß cells of the pancreas. One immunosuppressive agent that has recently shown promise in the treatment of new-onset T1D subjects aged 12-45 years is antithymocyte globulin (ATG), Thymoglobuline, encouraging further exploration in lower age groups. METHODS AND ANALYSIS: Minimal effective low dose (MELD)-ATG is a phase 2, multicentre, randomised, double-blind, placebo-controlled, multiarm parallel-group trial in participants 5-25 years diagnosed with T1D within 3-9 weeks of planned treatment day 1. A total of 114 participants will be recruited sequentially into seven different cohorts with the first cohort of 30 participants being randomised to placebo, 2.5 mg/kg, 1.5 mg/kg, 0.5 mg/kg and 0.1 mg/kg ATG total dose in a 1:1:1:1:1 allocation ratio. The next six cohorts of 12-15 participants will be randomised to placebo, 2.5 mg/kg, and one or two selected middle ATG total doses in a 1:1:1:1 or 1:1:1 allocation ratio, as dependent on the number of middle doses, given intravenously over two consecutive days. The primary objective will be to determine the changes in stimulated C-peptide response over the first 2 hours of a mixed meal tolerance test at 12 months for 2.5 mg/kg ATG arm vs the placebo. Conditional on finding a significant difference at 2.5 mg/kg, a minimally effective dose will be sought. Secondary objectives include the determination of the effects of a particular ATG treatment dose on (1) stimulated C-peptide, (2) glycated haemoglobin, (3) daily insulin dose, (4) time in range by intermittent continuous glucose monitoring measures, (5) fasting and stimulated dry blood spot (DBS) C-peptide measurements. ETHICS AND DISSEMINATION: MELD-ATG received first regulatory and ethical approvals in Belgium in September 2020 and from the German and UK regulators as of February 2021. The publication policy is set in the INNODIA (An innovative approach towards understanding and arresting Type 1 diabetes consortium) grant agreement (www.innodia.eu). TRIAL REGISTRATION NUMBER: NCT03936634; Pre-results.

Circulating biomarkers and outcomes from a randomised phase 2 trial of gemcitabine versus capecitabine-based chemoradiotherapy for pancreatic cancer.

BACKGROUND: The Phase 2 SCALOP trial compared gemcitabine with capecitabine-based consolidation chemoradiotherapy (CRT) in locally advanced pancreatic cancer (LAPC). METHODS: Thirty-five systematically identified circulating biomarkers were analysed in plasma samples from 60 patients enroled in SCALOP. Each was measured in triplicate at baseline (prior to three cycles of gemcitabine-capecitabine induction chemotherapy) and, for a subset, prior to CRT. Association with overall survival (OS) was determined using univariable Cox regression and optimal thresholds delineating low to high values identified using time-dependent ROC curves. Independence from known prognostic factors was assessed using Spearman correlation and the Wilcoxon rank sum test prior to multivariable Cox regression modelling including independent biomarkers and known prognostic factors. RESULTS: Baseline circulating levels of C-C motif chemokine ligand 5 (CCL5) were significantly associated with OS, independent of other clinicopathological characteristics. Patients with low circulating CCL5 (CCL5low) had a median OS of 18.5 (95% CI 11.76-21.32) months compared to 11.3 (95% CI 9.86-15.51) months in CCL5high; hazard ratio 1.95 (95% CI 1.04-8.65; p = 0.037). CONCLUSIONS: CCL5 is an independent prognostic biomarker in LAPC. Given the known role of CCL5 in tumour invasion, metastasis and the induction of an immunosuppressive micro-environment, targeting of CCL5-mediated pathways may offer therapeutic potential in pancreatic cancer. CLINICAL TRIAL REGISTRATION: The SCALOP trial was registered with ISRCTN, number 96169987 (registered 29 May 2008).

The Impact of Focality and Centricity on Vulvar Intraepithelial Neoplasia on Disease Progression in HIV+ Patients: A 10-Year Retrospective Study.

BACKGROUND: The impact of lesion focality and centricity in relation to patient outcome and disease recurrence of vulvar intraepithelial neoplasia (VIN) is an understudied area of research, especially in immunocompromised women. The prevalence and incidence of VIN have increased steadily since the 1980s because of the co-existence of human papillomavirus (HPV) and human immunodeficiency virus (HIV). In this study, we retrospectively examined the records of VIN patients to determine the effect of lesion focality and centricity with respect to the interval to disease recurrence. MATERIALS AND METHODS: All women diagnosed with VIN and managed between January 2002 and December 2011 were included (n = 90) and followed up until December 2017. Symptoms at the time of presentation, including HIV positivity (n = 75), were collated, including the influences of multifocality and multicentricity on time to disease recurrence. RESULTS: Multicentricity caused a more rapid recurrence of disease than unicentricity (p = 0.006), whereas multifocality increased the risk of recurrence more than unifocality (p < 0.0001). Viral load in the HIV+ patients was not associated with time to disease recurrence, but the reduced number of CD4+ lymphocytes present in HIV+ patients was. Treatment modalities had no effect on disease recurrence. CONCLUSION: Both focality and centricity have effects on interval to recurrence and final patient outcome, with multifocal disease having a poorer prognosis. Centricity and focality should be recorded at the time of diagnosis and act as a warning for disease recurrence. HIV+ VIN patients with multifocal disease and/or known immunosuppression (low CD4+ lymphocyte counts) should be regarded as "high-risk" patients and treated accordingly.

Genes Predisposed to DNA Hypermethylation during Acquired Resistance to Chemotherapy Are Identified in Ovarian Tumors by Bivalent Chromatin Domains at Initial Diagnosis.

Bivalent chromatin domains containing both active H3K4me3 and repressive H3K27me3 histone marks define gene sets poised for expression or silencing in differentiating embryonic stem (ES) cells. In cancer cells, aberrantly poised genes may facilitate changes in transcriptional states after exposure to anticancer drugs. In this study, we used ChIP-seq to characterize genome-wide positioning of H3K4me3- and H3K27me3-associated chromatin in primary high-grade serous ovarian carcinomas and in normal ovarian surface and fallopian tube tissue. Gene sets with proximal bivalent marks defined in this manner were evaluated subsequently as signatures of systematic change in DNA methylation and gene expression, comparing pairs of tissue samples taken from patients at primary presentation and relapse following chemotherapy. We found that gene sets harboring bivalent chromatin domains at their promoters in tumor tissue, but not normal epithelia, overlapped with Polycomb-repressive complex target genes as well as transcriptionally silenced genes in normal ovarian and tubal stem cells. The bivalently marked genes we identified in tumors before chemotherapy displayed increased promoter CpG methylation and reduced gene expression at relapse after chemotherapy of ovarian cancer. Overall, our results support the hypothesis that preexisting histone modifications at genes in a poised chromatin state may lead to epigenetic silencing during acquired drug resistance.Significance: These results suggest epigenetic targets for intervention to prevent the emergence of cancer drug resistance. Cancer Res; 78(6); 1383-91. ©2018 AACR.

NY-ESO-1 expression in DCIS: A new predictor of good prognosis.

BACKGROUND: At present, it is difficult to predict which patients with ductal carcinoma-in-situ (DCIS) will subsequently develop frank invasive breast cancer (IDC). A recent survey by our group has shown that NY-ESO-1 and MAGEA are both expressed in DCIS. This study was aimed at determining whether expression of these antigens was related to the later development of IDC. RESULTS: 14 of 42 (33%) of patients developed invasive breast cancer during the follow up period. Only one of those DCIS cases that relapsed was positive for NYESO-1 at diagnosis. In contrast, DCIS samples of 15 of the 28 (54%) of those patients who remained disease-free expressed NY-ESO-1. (Permutation chi square p=0.0033). METHODS: We identified 42 patients with DCIS, and followed them up for more than 10 years. NY-ESO-1 and MAGEA were demonstrated by immunostaining as were CD8+ infiltrates on all sections together with the conventional markers, ER, PR, and HER2. CONCLUSIONS: Expression of NY-ESO-1 may predict those patients who will not subsequently develop invasive breast cancer and could therefore potentially be helpful in defining prognosis in patients with DCIS.

Methylation of MYLK3 gene promoter region: a biomarker to stratify surgical care in ovarian cancer in a multicentre study.

BACKGROUND: Survival benefit from surgical debulking of ovarian cancer (OC) is well established, but some women, despite total macroscopic clearance of disease, still have poor prognosis. We aimed to identify biomarkers to predict benefit from conventional surgery. METHODS: Clinical data from women debulked for high-stage OC were analysed (Hammersmith Hospital, London, UK; 2001-2014). Infinium's HumanMethylation27 array interrogated tumour DNA for differentially methylated CpG sites, correlated to survival, in patients with the least residual disease (RD; Hammersmith Array). Validation was performed using bisulphite pyrosequencing (Charité Hospital, Berlin, Germany cohort) and The Cancer Genome Atlas' (TCGA) methylation data set. Kaplan-Meier curves and Cox models tested survival. RESULTS: Altogether 803 women with serous OC were studied. No RD was associated with significantly improved overall survival (OS; hazard ratio (HR) 1.25, 95% CI 1.06-1.47; P=0.0076) and progression-free survival (PFS; HR 1.23, 95% CI 1.05-1.43; P=0.012; Hammersmith database n=430). Differentially methylated loci within FGF4, FGF21, MYLK2, MYLK3, MYL7, and ITGAE associated with survival. Patients with the least RD had significantly better OS with higher methylation of MYLK3 (Hammersmith (HR 0.51, 95% CI 0.31-0.84; P=0.01), Charité (HR 0.46, 95% CI 0.21-1.01; P=0.05), and TCGA (HR 0.64, 95% CI 0.44-0.93; P=0.02)). CONCLUSIONS: MYLK3 methylation is associated with improved OS in patients with the least RD, which could potentially be used to determine response to surgery.

Challenges and methodology in the incorporation of biomarkers in cancer clinical trials.

Biomarkers can be used to establish more homogeneous groups using the genetic makeup of the tumour to inform the selection of treatment for each individual patient. However, proper preclinical work and stringent validation are needed before taking forward biomarkers into confirmatory studies. Despite the challenges, incorporation of biomarkers into clinical trials could better target appropriate patients, and potentially be lifesaving. The authors conducted a systematic review to describe marker-based and adaptive design methodology for their integration in clinical trials, and to further describe the associated practical challenges. Studies published between 1990 to November 2015 were searched on PubMed. Titles, abstracts and full text articles were reviewed to identify relevant studies. Of the 4438 studies examined, 57 studies were included. The authors conclude that the proposed approaches may readily help researchers to design biomarker trials, but novel approaches are still needed.

Gestational length assignment based on last menstrual period, first trimester crown-rump length, ovulation, and implantation timing.

PURPOSE: Understanding the natural length of human pregnancy is central to clinical care. However, variability in the reference methods to assign gestational age (GA) confound our understanding of pregnancy length. Assignation from ultrasound measurement of fetal crown-rump length (CRL) has superseded that based on last menstrual period (LMP). Our aim was to estimate gestational length based on LMP, ultrasound CRL, and implantation that were known, compared to pregnancy duration assigned by day of ovulation. METHODS: Prospective study in 143 women trying to conceive. In 71 ongoing pregnancies, gestational length was estimated from LMP, CRL at 10-14 weeks, ovulation, and implantation day. For each method of GA assignment, the distribution in observed gestational length was derived and both agreement and correlation between the methods determined. RESULTS: Median ovulation and implantation days were 16 and 27, respectively. The gestational length based on LMP, CRL, implantation, and ovulation was similar: 279, 278, 276.5 and 276.5 days, respectively. The distributions for observed gestational length were widest where GA was assigned from CRL and LMP and narrowest when assigned from implantation and ovulation day. The strongest correlation for gestational length assessment was between ovulation and implantation (r = 0.98) and weakest between CRL and LMP (r = 0.88). CONCLUSIONS: The most accurate method of predicting gestational length is ovulation day, and this agrees closely with implantation day. Prediction of gestational length from CRL and known LMP are both inferior to ovulation and implantation day. This information could have important implications on the routine assignment of gestational age.

Epigenetic Regulation of the Homeobox Gene MSX1 Associates with Platinum-Resistant Disease in High-Grade Serous Epithelial Ovarian Cancer.

PURPOSE: Although high-grade serous ovarian cancer (HGSOC) is frequently chemoresponsive, a proportion of patients do not respond to platinum-based chemotherapy at presentation or have progression-free survival (PFS) of less than 6 months. Validated predictive biomarkers of lack of response would enable alternative treatment stratification for these patients and identify novel mechanisms of intrinsic resistance. Our aim was to identify DNA methylation biomarkers of poor response to chemotherapy and demonstrate involvement of the associated gene in platinum drug cell sensitivity. EXPERIMENTAL DESIGN: DNA methylation was investigated in independent tumor cohorts using Illumina HumanMethylation arrays and gene expression by Affymetrix arrays and qRT-PCR. The role of Msh homeobox 1 (MSX1) in drug sensitivity was investigated by gene reintroduction and siRNA knockdown of ovarian cancer cell lines. RESULTS: CpG sites at contiguous genomic locations within the MSX1 gene have significantly lower levels of methylation in independent cohorts of HGSOC patients, which recur by 6 months compared with after 12 months (P < 0.05, q < 0.05, n = 78), have poor RECIST response (P < 0.05, q < 0.05, n = 61), and are associated with PFS in an independent cohort (n = 146). A decrease in methylation at these CpG sites correlates with decreased MSX1 gene expression. MSX1 expression is associated with PFS (HR, 0.92; 95% CI, 0.85-0.99; P = 0.029; n = 309). Cisplatin-resistant ovarian cancer cell lines have reduced MSX1 expression, and MSX1 overexpression leads to cisplatin sensitization, increased apoptosis, and increased cisplatin-induced p21 expression. CONCLUSIONS: Hypomethylation of CpG sites within the MSX1 gene is associated with resistant HGSOC disease at presentation and identifies expression of MSX1 as conferring platinum drug sensitivity. Clin Cancer Res; 22(12); 3097-104. ©2016 AACR.

Transcriptional implications of intragenic DNA methylation in the oestrogen receptor alpha gene in breast cancer cells and tissues.

BACKGROUND: DNA methylation variability regions (MVRs) across the oestrogen receptor alpha (ESR1) gene have been identified in peripheral blood cells from breast cancer patients and healthy individuals. In contrast to promoter methylation, gene body methylation may be important in maintaining active transcription. This study aimed to assess MVRs in ESR1 in breast cancer cell lines, tumour biopsies and exfoliated epithelial cells from expressed breast milk (EBM), to determine their significance for ESR1 transcription. METHODS: DNA methylation levels in eight MVRs across ESR1 were assessed by pyrosequencing bisulphite-converted DNA from three oestrogen receptor (ER)-positive and three ER-negative breast cancer cell lines. DNA methylation and expression were assessed following treatment with DAC (1 µM), or DMSO (controls). ESR1 methylation levels were also assayed in DNA from 155 invasive ductal carcinoma biopsies provided by the Breast Cancer Campaign Tissue Bank, and validated with DNA methylation profiles from the TCGA breast tumours (n = 356 ER-pos, n = 109 ER-neg). DNA methylation was profiled in exfoliated breast epithelial cells from EBM using the Illumina 450 K (n = 36) and pyrosequencing in a further 53 donor samples. ESR1 mRNA levels were measured by qRT-PCR. RESULTS: We show that ER-positive cell lines had unmethylated ESR1 promoter regions and highly methylated intragenic regions (median, 80.45%) while ER-negative cells had methylated promoters and lower intragenic methylation levels (median, 38.62%). DAC treatment increased ESR1 expression in ER-negative cells, but significantly reduced methylation and expression of ESR1 in ER-positive cells. The ESR1 promoter was unmethylated in breast tumour biopsies with high levels of intragenic methylation, independent of ER status. However, ESR1 methylation in the strongly ER-positive EBM DNA samples were very similar to ER-positive tumour cell lines. CONCLUSION: DAC treatment inhibited ESR1 transcription in cells with an unmethylated ESR1 promoter and reduced intragenic DNA methylation. Intragenic methylation levels correlated with ESR1 expression in homogenous cell populations (cell lines and exfoliated primary breast epithelial cells), but not in heterogeneous tumour biopsies, highlighting the significant differences between the in vivo tumour microenvironment and individual homogenous cell types. These findings emphasise the need for care when choosing material for epigenetic research and highlights the presence of aberrant intragenic methylation levels in tumour tissue.

Poised epigenetic states and acquired drug resistance in cancer.

Epigenetic events, which are somatically inherited through cell division, are potential drivers of acquired drug resistance in cancer. The high rate of epigenetic change in tumours generates diversity in gene expression patterns that can rapidly evolve through drug selection during treatment, leading to the development of acquired resistance. This will potentially confound stratified chemotherapy decisions that are solely based on mutation biomarkers. Poised epigenetic states in tumour cells may drive multistep epigenetic fixation of gene expression during the acquisition of drug resistance, which has implications for clinical strategies to prevent the emergence of drug resistance.

The prognostic significance of the aberrant extremes of p53 immunophenotypes in breast cancer.

AIMS: The utility of p53 as a prognostic assay has been elusive. The aims of this study were to describe a novel, reproducible scoring system and assess the relationship between differential p53 immunohistochemistry (IHC) expression patterns, TP53 mutation status and patient outcomes in breast cancer. METHODS AND RESULTS: Tissue microarrays were used to study p53 IHC expression patterns: expression was defined as extreme positive (EP), extreme negative (EN), and non-extreme (NE; intermediate patterns). Overall survival (OS) was used to define patient outcome. A representative subgroup (n = 30) showing the various p53 immunophenotypes was analysed for TP53 hotspot mutation status (exons 4-9). Extreme expression of any type occurred in 176 of 288 (61%) cases. As compared with NE expression, EP expression was significantly associated (P = 0.039) with poorer OS. In addition, as compared with NE expression, EN expression was associated (P = 0.059) with poorer OS. Combining cases showing either EP or EN expression better predicted OS than either pattern alone (P = 0.028). This combination immunophenotype was significant in univariate but not multivariate analysis. In subgroup analysis, six substitution exon mutations were detected, all corresponding to extreme IHC phenotypes. Five missense mutations corresponded to EP staining, and the nonsense mutation corresponded to EN staining. No mutations were detected in the NE group. CONCLUSIONS: Patients with extreme p53 IHC expression have a worse OS than those with NE expression. Accounting for EN as well as EP expression improves the prognostic impact. Extreme expression positively correlates with nodal stage and histological grade, and negatively with hormone receptor status. Extreme expression may relate to specific mutational status.

Plasma antibodies to oral bacteria and risk of pancreatic cancer in a large European prospective cohort study.

OBJECTIVE: Examine the relationship between antibodies to 25 oral bacteria and pancreatic cancer risk in a prospective cohort study. DESIGN: We measured antibodies to oral bacteria in prediagnosis blood samples from 405 pancreatic cancer cases and 416 matched controls, nested within the European Prospective Investigation into Cancer and Nutrition study. Analyses were conducted using conditional logistic regression and additionally adjusted for smoking status and body mass index. RESULTS: Individuals with high levels of antibodies against Porphyromonas gingivalis ATTC 53978, a pathogenic periodontal bacteria, had a twofold higher risk of pancreatic cancer than individuals with lower levels of these antibodies (OR 2.14; 95% CI 1.05 to 4.36; >200 ng/ml vs ≤200 ng/ml). To explore the association with commensal (non-pathogenic) oral bacteria, we performed a cluster analysis and identified two groups of individuals, based on their antibody profiles. A cluster with overall higher levels of antibodies had a 45% lower risk of pancreatic cancer than a cluster with overall lower levels of antibodies (OR 0.55; 95% CI 0.36 to 0.83). CONCLUSIONS: Periodontal disease might increase the risk for pancreatic cancer. Moreover, increased levels of antibodies against specific commensal oral bacteria, which can inhibit growth of pathogenic bacteria, might reduce the risk of pancreatic cancer. Studies are needed to determine whether oral bacteria have direct effects on pancreatic cancer pathogenesis or serve as markers of the immune response.

Ambient particulate matter and the response to orthostatic challenge in the elderly: the Maintenance of Balance, Independent Living, Intellect, and Zest in the Elderly (MOBILIZE) of Boston study.

Short-term elevations in ambient fine particulate matter (PM(2.5)) may increase resting systolic (SBP) and diastolic (DBP) blood pressures, but whether PM(2.5) alters hemodynamic responses to orthostatic challenge has not been studied in detail. We repeatedly measured SBP and DBP during supine rest and 1 and 3 minutes after standing among 747 elderly (aged 78.3±5.3 years, mean±SD) participants from a prospective cohort study. We used linear mixed models to assess the association between change in SBP (ΔSBP=standing SBP-supine SBP) and DBP (ΔDBP) on standing and mean PM(2.5) levels over the preceding 1 to 28 days, adjusting for meteorologic covariates, temporal trends, and medical history. We observed a 1.4-mm Hg (95% CI: 0.0-2.8 mm Hg; P=0.046) higher ΔSBP and a 0.7-mm Hg (95% CI: 0.0-1.4 mm Hg; P=0.053) higher ΔDBP at 1 minute of standing per interquartile range increase (3.8 µg/m(3)) in mean PM(2.5) levels in the past 7 days. ΔSBP and ΔDBP measured 3 minutes after standing were not associated with PM(2.5). Resting DBP (but not SBP or pulse pressure) was positively associated with PM(2.5) at longer averaging periods. Responses were more strongly associated with black carbon than sulfate levels. These associations did not differ significantly according to hypertension status, obesity, diabetes mellitus, or sex. These results suggest that ambient particles can increase resting DBP and exaggerate blood pressure responses to postural changes in elderly people. Increased vasoreactivity during posture change may be responsible, in part, for the adverse effect of ambient particles on cardiovascular health.

In utero exposures, infant growth, and DNA methylation of repetitive elements and developmentally related genes in human placenta.

BACKGROUND: Fetal programming describes the theory linking environmental conditions during embryonic and fetal development with risk of diseases later in life. Environmental insults in utero may lead to changes in epigenetic mechanisms potentially affecting fetal development. OBJECTIVES: We examined associations between in utero exposures, infant growth, and methylation of repetitive elements and gene-associated DNA in human term placenta tissue samples. METHODS: Placental tissues and associated demographic and clinical data were obtained from subjects delivering at Women and Infants Hospital in Providence, Rhode Island (USA). Methylation levels of long interspersed nuclear element-1 (LINE-1) and the Alu element AluYb8 were determined in 380 placental samples from term deliveries using bisulfite pyrosequencing. Genomewide DNA methylation profiles were obtained in a subset of 184 samples using the Illumina Infinium HumanMethylation27 BeadArray. Multiple linear regression, model-based clustering methods, and gene set enrichment analysis examined the association between birth weight percentile, demographic variables, and repetitive element methylation and gene-associated CpG locus methylation. RESULTS: LINE-1 and AluYb8 methylation levels were found to be significantly positively associated with birth weight percentile (p = 0.01 and p < 0.0001, respectively) and were found to differ significantly among infants exposed to tobacco smoke and alcohol. Increased placental AluYb8 methylation was positively associated with average methylation among CpG loci found in polycomb group target genes; developmentally related transcription factor binding sites were overrepresented for differentially methylated loci associated with both elements. CONCLUSIONS: Our results suggest that repetitive element methylation markers, most notably AluYb8 methylation, may be susceptible to epigenetic alterations resulting from the intrauterine environment and play a critical role in mediating placenta function, and may ultimately inform on the developmental basis of health and disease.

Association of secondhand smoke exposures with DNA methylation in bladder carcinomas.

BACKGROUND: The association between secondhand smoke (SHS) exposure and bladder cancer is inconclusive. Epigenetic alterations in bladder tumors have been linked to primary cigarette smoking and could add to the biological plausibility of an association between SHS exposure and bladder cancer. HYPOTHESIS: SHS exposure is associated with DNA methylation in bladder tumors. METHODS: Using an array-based approach, we profiled DNA methylation from never smoking cases of incident bladder cancer. Analyses examined associations between individual loci's methylation with SHS variables (exposure in adulthood, childhood, occupationally, and total exposure). A canonical pathway analysis was used to find pathways significantly associated with each SHS exposure type. RESULTS: There is a trend toward increased methylation of numerous CpG loci with increasing exposure to adulthood, occupational, and total SHS. Discrete associations between methylation extent of several CpG loci and SHS exposures demonstrated significantly increased methylation of these loci across all types of SHS exposure. CpGs with SHS-related methylation alterations were associated with genes in pathways involved in carcinogenesis, immune modulation, and immune signaling. INTERPRETATION: Exposures to SHS in adulthood, childhood, occupationally, and in total are each significantly associated with changes in DNA methylation of several CpG loci in bladder tumors, adding biological plausibility to SHS as a risk factor for bladder cancer.