DNA methylation instability by BRAF-mediated TET silencing and lifestyle-exposure divides colon cancer pathways.

BACKGROUND: Aberrations in DNA methylation are widespread in colon cancer (CC). Understanding origin and progression of DNA methylation aberrations is essential to develop effective preventive and therapeutic strategies. Here, we aimed to dissect CC subtype-specific methylation instability to understand underlying mechanisms and functions. METHODS: We have assessed genome-wide DNA methylation in the healthy normal colon mucosa (HNM), precursor lesions and CCs in a first comprehensive study to delineate epigenetic change along the process of colon carcinogenesis. Mechanistically, we used stable cell lines, genetically engineered mouse model of mutant BRAFV600E and molecular biology analysis to establish the role of BRAFV600E-mediated-TET inhibition in CpG-island methylator phenotype (CIMP) inititation. RESULTS: We identified two distinct patterns of CpG methylation instability, determined either by age-lifestyle (CC-neutral CpGs) or genetically (CIMP-CpGs). CC-neutral-CpGs showed age-dependent hypermethylation in HNM, all precursors, and CCs, while CIMP-CpGs showed hypermethylation specifically in sessile serrated adenomas/polyps (SSA/Ps) and CIMP-CCs. BRAFV600E-mutated CCs and precursors showed a significant downregulation of TET1 and TET2 DNA demethylases. Stable expression of BRAFV600E in nonCIMP CC cells and in a genetic mouse model was sufficient to repress TET1/TET2 and initiate hypermethylation at CIMP-CpGs, reversible by BRAFV600E inhibition. BRAFV600E-driven CIMP-CpG hypermethylation occurred at genes associated with established CC pathways, effecting functional changes otherwise achieved by genetic mutation in carcinogenesis. CONCLUSIONS: Hence, while age-lifestyle-driven hypermethylation occurs generally in colon carcinogenesis, BRAFV600E-driven hypermethylation is specific for the "serrated" pathway. This knowledge will advance the use of epigenetic biomarkers to assess subgroup-specific CC risk and disease progression.

Tumor Initiation Capacity and Therapy Resistance Are Differential Features of EMT-Related Subpopulations in the NSCLC Cell Line A549.

Cell lines are essential tools to standardize and compare experimental findings in basic and translational cancer research. The current dogma states that cancer stem cells feature an increased tumor initiation capacity and are also chemoresistant. Here, we identified and comprehensively characterized three morphologically distinct cellular subtypes in the non-small cell lung cancer cell line A549 and challenge the current cancer stem cell dogma. Subtype-specific cellular morphology is maintained during short-term culturing, resulting in the formation of holoclonal, meroclonal, and paraclonal colonies. A549 holoclone cells were characterized by an epithelial and stem-like phenotype, paraclone cells featured a mesenchymal phenotype, whereas meroclone cells were phenotypically intermediate. Cell-surface marker expression of subpopulations changed over time, indicating an active epithelial-to-mesenchymal transition (EMT), in vitro and in vivo. EMT has been associated with the overexpression of the immunomodulators PD-L1 and PD-L2, which were 37- and 235-fold overexpressed in para- versus holoclone cells, respectively. We found that DNA methylation is involved in epigenetic regulation of marker expression. Holoclone cells were extremely sensitive to cisplatin and radiotherapy in vitro, whereas paraclone cells were highly resistant. However, inhibition of the receptor tyrosine kinase AXL, whose expression is associated with an EMT, specifically targeted the otherwise highly resistant paraclone cells. Xenograft tumor formation capacity was 24- and 269-fold higher in holo- than mero- and paraclone cells, respectively. Our results show that A549 subpopulations might serve as a unique system to explore the network of stemness, cellular plasticity, tumor initiation capacity, invasive and metastatic potential, and chemo/radiotherapy resistance.

Modulation of age- and cancer-associated DNA methylation change in the healthy colon by aspirin and lifestyle.

BACKGROUND: Aberrant DNA methylation in gene promoters is associated with aging and cancer, but the circumstances determining methylation change are unknown. We investigated the impact of lifestyle modulators of colorectal cancer (CRC) risk on the stability of gene promoter methylation in the colonic mucosa. METHODS: We measured genome-wide promoter CpG methylation in normal colon biopsies (n = 1092) from a female screening cohort, investigated the interaction of lifestyle factors with age-dependent increase in methylation with log-linear multivariable regression, and related their modifying effect to hypermethylation in CRC. All statistical tests were two-sided. RESULTS: Of 20025 promoter-associated CpGs analyzed, 1713 showed statistically significant age-dependent methylation gains. Fewer CpGs acquired methylation in users of aspirin (≥ 2 years) and hormonal replacement therapy (HRT age ≥ 50 years) compared with nonusers (43 vs 1355; 1 vs1377, respectively), whereas more CpGs were affected in smokers (≥ 20 years) and individuals with a body mass index (BMI) of 25 kg/m(2) and greater compared with control groups (180 vs 39; 554 vs 144, respectively). Fifty percent of the CpGs showing age-dependent methylation were found hypermethylated in CRC (odds ratio [OR] = 20; 95% confidence interval [CI] = 18 to 23; P < 2 × 10(-16)). These loci gained methylation with a higher median rate compared with age-only methylated sites (P = 2 × 10(-76)) and were enriched for polycomb regions (OR = 3.67). Importantly, aspirin (P < .001) and HRT use (P < .001) reduced the methylation rate at these cancer-related genes, whereas smoking (P < .001) and high BMI (P = .004) increased it. CONCLUSIONS: Lifestyle, including aspirin use, modulates age-associated DNA methylation change in the colonic epithelium and thereby impacts the evolution of cancer methylomes.

Optomechanically induced transparency.

Electromagnetically induced transparency is a quantum interference effect observed in atoms and molecules, in which the optical response of an atomic medium is controlled by an electromagnetic field. We demonstrated a form of induced transparency enabled by radiation-pressure coupling of an optical and a mechanical mode. A control optical beam tuned to a sideband transition of a micro-optomechanical system leads to destructive interference for the excitation of an intracavity probe field, inducing a tunable transparency window for the probe beam. Optomechanically induced transparency may be used for slowing and on-chip storage of light pulses via microfabricated optomechanical arrays.

Blunted cortisol responses to psychosocial stress in asthmatic children: a general feature of atopic disease?

OBJECTIVE: Atopy is defined by the individual predisposition to develop a group of inflammatory disorders in response to certain food or environmental substances that are otherwise innocuous for the host. In previous studies we could demonstrate a reduced responsiveness of the hypothalamus-pituitary-adrenal (HPA) axis to psychosocial stress in young and adult patients with atopic dermatitis (AD), a chronic atopic skin disorder. With respect to the important immunoregulatory role of the HPA axis, especially under stress, this observation could be of clinical relevance and may at least partly explain stress-induced exacerbation of AD. The present study was designed to investigate whether attenuated responsiveness of the HPA axis to stress represents a characteristic feature of AD or whether it can also be found in other chronic manifestations of atopy. METHODS: Children (aged 7-12) with allergic asthma (AA; N = 17) and age- and sex-matched healthy controls (N = 18) were exposed to the "Trier Social Stress Test for Children"(TSST-C), which mainly consists of a free speech and mental arithmetic tasks in front of an audience. Salivary cortisol was measured in ten-minute intervals before and after the TSST-C, while heart rate was monitored continuously. In addition, early morning cortisol levels (after awakening, +10, +20, +30 minutes) were assessed on three consecutive days. RESULTS: Data analysis yielded a significant increase of cortisol concentrations (F (9297)= 16.79; p <.001) and heart rates (F(32,992)= 9.16; p <.001) after the stressor with no between-group difference in heart rate responses. However, AA children showed a significantly blunted cortisol response to the TSST-C when compared with the control group (F(9297)= 2.95; p <.01). Awakening in the morning was accompanied by a significant rise of cortisol levels on all three experimental days in AA and control subjects (all p <.001) that was not different between the two groups. CONCLUSIONS: These findings suggest that a blunted adrenocortical response to stress may represent a common feature of chronic allergic inflammatory processes that may be relevant in different forms of chronic manifestation of atopy.