Long-range epigenetic silencing at 2q14.2 affects most human colorectal cancers and may have application as a non-invasive biomarker of disease

Large chromosomal regions can be suppressed in cancer cells as denoted by hypermethylation of neighbouring CpG islands and downregulation of most genes within the region. We have analysed the extent and prevalence of long-range epigenetic silencing at 2q14.2 (the first and best characterised example of coordinated epigenetic remodelling) and investigated its possible applicability as a non-invasive diagnostic marker of human colorectal cancer using different approaches and biological samples. Hypermethylation of at least one of the CpG islands analysed (EN1, SCTR, INHBB) occurred in most carcinomas (90%), with EN1 methylated in 73 and 40% of carcinomas and adenomas, respectively. Gene suppression was a common phenomenon in all the tumours analysed and affected both methylated and unmethylated genes. Detection of methylated EN1 using bisulfite treatment and melting curve (MC) analysis from stool DNA in patients and controls resulted in a predictive capacity of, 44% sensitivity in positive patients (27% of overall sensitivity) and 97% specificity. We conclude that epigenetic suppression along 2q14.2 is common to most colorectal cancers and the presence of a methylated EN1 CpG island in stool DNA might be used as biomarker of neoplastic disease

The aberrant asynchronous replication — characterizing lymphocytes of cancer patients — is erased following stem cell transplantation

<p>Abstract</p> <p>Background</p> <p>Aberrations of allelic replication timing are epigenetic markers observed in peripheral blood cells of cancer patients. The aberrant markers are non-cancer-type-specific and are accompanied by increased levels of sporadic aneuploidy. The study aimed at following the epigenetic markers and aneuploidy levels in cells of patients with haematological malignancies from diagnosis to full remission, as achieved by allogeneic stem cell transplantation (alloSCT).</p> <p>Methods</p> <p><it>TP53 </it>(a tumor suppressor gene assigned to chromosome 17), <it>AML1 </it>(a gene assigned to chromosome 21 and involved in the leukaemia-abundant 8;21 translocation) and the pericentomeric satellite sequence of chromosome 17 (<it>CEN17</it>) were used for replication timing assessments. Aneuploidy was monitored by enumerating the copy numbers of chromosomes 17 and 21. Replication timing and aneuploidy were detected cytogenetically using fluorescence <it>in situ </it>hybridization (FISH) technology applied to phytohemagglutinin (PHA)-stimulated lymphocytes.</p> <p>Results</p> <p>We show that aberrant epigenetic markers are detected in patients with hematological malignancies from the time of diagnosis through to when they are scheduled to undergo alloSCT. These aberrations are unaffected by the clinical status of the disease and are displayed both during accelerated stages as well as in remission. Yet, these markers are eradicated completely following stem cell transplantation. In contrast, the increased levels of aneuploidy (irreversible genetic alterations) displayed in blood lymphocytes at various stages of disease are not eliminated following transplantation. However, they do not elevate and remain unchanged (stable state). A demethylating anti-cancer drug, 5-azacytidine, applied in vitro to lymphocytes of patients prior to transplantation mimics the effect of transplantation: the epigenetic aberrations disappear while aneuploidy stays unchanged.</p> <p>Conclusions</p> <p>The reversible nature of the replication aberrations may serve as potential epigenetic blood markers for evaluating the success of transplant or other treatments and for long-term follow up of the patients who have overcome a hematological malignancy.</p

Pαx6 Expression in Postmitotic Neurons Mediates the Growth of Axons in Response to SFRP1

During development, the mechanisms that specify neuronal subclasses are coupled to those that determine their axonal response to guidance cues. Pax6 is a homedomain transcription factor required for the specification of a variety of neural precursors. After cell cycle exit, Pax6 expression is often shut down in the precursor progeny and most postmitotic neurons no longer express detectable levels of the protein. There are however exceptions and high Pax6 protein levels are found, for example, in postmitotic retinal ganglion cells (RGCs), dopaminergic neurons of the olfactory bulb and the limbic system in the telencephalon. The function of Pax6 in these differentiating neurons remains mostly elusive. Here, we demonstrate that Pax6 mediates the response of growing axons to SFRP1, a secreted molecule expressed in several Pax6-positive forebrain territories. Forced expression of Pax6 in cultured postmitotic cortical neurons, which do not normally express Pax6, was sufficient to increment axonal length. Growth was blocked by the addition of anti-SFRP1 antibodies, whereas exogenously added SFRP1 increased axonal growth of Pax6-transfected neurons but not that of control or untransfected cortical neurons. In the reverse scenario, shRNA-mediated knock-down of Pax6 in mouse retinal explants specifically abolished RGCs axonal growth induced by SFRP1, but had no effect on RGCs differentiation and it did not modify the effect of Shh or Netrin on axon growth. Taken together these results demonstrate that expression of Pax6 is necessary and sufficient to render postmitotic neurons competent to respond to SFRP1. These results reveal a novel and unexpected function of Pax6 in postmitotic neurons and situate Pax6 and SFRP1 as pair regulators of axonal connectivity

Low rates of mutation in clinical grade human pluripotent stem cells under different culture conditions

Abstract: The occurrence of repetitive genomic changes that provide a selective growth advantage in pluripotent stem cells is of concern for their clinical application. However, the effect of different culture conditions on the underlying mutation rate is unknown. Here we show that the mutation rate in two human embryonic stem cell lines derived and banked for clinical application is low and not substantially affected by culture with Rho Kinase inhibitor, commonly used in their routine maintenance. However, the mutation rate is reduced by >50% in cells cultured under 5% oxygen, when we also found alterations in imprint methylation and reversible DNA hypomethylation. Mutations are evenly distributed across the chromosomes, except for a slight increase on the X-chromosome, and an elevation in intergenic regions suggesting that chromatin structure may affect mutation rate. Overall the results suggest that pluripotent stem cells are not subject to unusually high rates of genetic or epigenetic alterations

Gene switching rate determines response to extrinsic perturbations in the self-activation transcriptional network motif

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Modifications of Microvascular EC Surface Modulate Phototoxicity of a Porphycene anti-ICAM-1 Immunoconjugate; Therapeutic Implications

Inflammation and shear stress can upregulate expression of cellular adhesion molecules in endothelial cells (EC). The modified EC surface becomes a mediating interface between the circulating blood elements and the endothelium, and grants opportunity for immunotherapy. In photodynamic therapy (PDT), immunotargeting might overcome the lack of selectivity of currently used sensitizers. In this study, we hypothesized that differential ICAM-1 expression modulates the effects of a drug targeted to surface ICAM-1. A novel porphycene–anti-ICAM-1 conjugate was synthesized and applied to treat endothelial cells from macro and microvasculature. Results show that the conjugate induces phototoxicity in inflamed, but not in healthy, microvascular EC. Conversely, macrovascular EC exhibited phototoxicity regardless of their state. These findings have two major implications; the relevance of ICAM-1 as a modulator of drug effects in microvasculature, and the potential of the porphycene bioconjugate as a promising novel PDT agent.Spain. Ministerio de Economia y Competividad (Grant BFU2009-0984)Spain. Ministerio de Economia y Competividad (Grant CTQ2010-20870-C03-01)National Institutes of Health (U.S.) (Grant NIH/NIGMS RO1/GM049039)Fundacio Empreses IQSFundacio Empreses POSIMATMIT-Spain Seed Fun