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Diffuse intrinsic pontine gliomas (DIPGs) are lethal brain tumors with no effective therapies other than radiation. Inhibitor of DNA binding (ID) proteins are key regulators of tissue and lineage-specific gene differentiation during embryogenesis. Previous work has shown that H3F3A and ACVR1 mutations increase ID1 expression in cultured astrocytes, but this has not been validated in human DIPG, nor has the regulation and targetability of ID1 been explored in DIPG. Analysis of multi-focal post-mortem tumor and normal brain tissue (n=52) as well as multiple human datasets revealed ID1 to be elevated in DIPG. Elevated ID1 was found to correlate with reduced survival in DIPG. In a multi-focal autopsy DIPG case, we found ID1 expression to be heterogeneous and to correlate with tumor invasion. Chromatin immunoprecipitation-qPCR (ChIP-qPCR) was used to quantify H3K27ac and H3K27me3 at ID1 gene regulatory regions (promoters/enhancers) in multi-focal post-mortem tissue. The ID1 loci was found to be epigenetically poised for up-regulation (elevated H3K27ac and low H3K27me3) in DIPG tissue, regardless of H3 or ACVR1 mutation status, compared to normal brain. Analysis of publically-available ISH and ChIP-sequencing data revealed elevated ID1 expression and ID1-enhancer H3K27ac in prenatal mouse hindbrain compared to prenatal forebrain, prenatal midbrain, and all postnatal brain regions. ID1 shRNA-mediated knockdown of primary human H3K27M DIPG cells (DIPG007) resulted in significantly reduced invasion and migration. As cannabidiol (CBD) has successfully been used to therapeutically target ID1 in pre-clinical models of adult human cancers, we treated DIPG007 cells with CBD and found reduced viability at clinically relevant dosing (IC50=2.4 uM) with dose-dependent reduction in ID1 protein. ID1 knockdown and CBD treatment studies in murine models of DIPG are ongoing. These findings indicate that a multifactorial (genetic and regionally-based) epigenetic upregulation of ID1 drives DIPG invasiveness and is targetable with CBD.

Nearby Type III (galaxy-spanning) Kardashev supercivilizations would have high mid-infrared (MIR) luminosities. We have used the Wide-field Infrared Survey Explorer (WISE) to survey ~$1 \times 10^5$ galaxies for extreme MIR emission, $10^3$ times more galaxies than the only previous such search. We have calibrated the WISE All-sky Catalog pipeline products to improve its photometry for extended sources. We present 563 extended sources with $|b| \ge 10$ and red MIR colors, having visually vetted them to remove artifacts. No galaxies in our sample host an alien civilization reprocessing more than 85% of its starlight into the MIR, and only 50 galaxies, including Arp 220, have MIR luminosities consistent with >50% reprocessing. Ninety of these (likely) extragalactic sources have little literature presence; in most cases they are likely barely resolved galaxies or pairs of galaxies undergoing large amounts of star formation. Five are new to science and deserve further study. The Be star 48 Librae sits within a MIR nebula, and we suggest that it may be creating dust. WISE, 2MASS, and Spitzer imagery shows that IRAS 04287+6444 is consistent with a previously unnoticed, heavily extinguished cluster of young stellar objects. We identify five "passive" (i.e. red) spiral galaxies with unusually high MIR and low NUV luminosity. We search a set of optically "dark" HI galaxies for MIR emission, and find none. These 90 poorly understood sources and five anomalous passive spirals deserve follow-up via both SETI and conventional astrophysics., ApJS 217 25. 44pp, 9 tables, 25 figures. Due to limitations of arXiv LaTeX compilation, figures are sub-optimally placed and tables are not landscape. See http://astro.psu.edu/~jtwright/Dyson/GHAT3.pdf for version with full resolution figures and fully legible tables

Figure S1. Serial dilution of K27M mutant oligonucleotide in constant background of wild-type DNA demonstrates consistent detection down to at least 2% VAF under typical experimental conditions, with the possibility of detection at even lower VAF under ideal conditions. One such dilution series is shown above, with (a) showing number of droplets positive for mutant or wild-type H3F3A sequence and (b) showing the corresponding VAF values. Figure S2. Plot of droplets (blue â positive mutant H3F3A K27M, green â positive wildtype H3F3A, grey â negative droplets) from ddPCR performed on (a) non-tumor human CSF spiked with synthetic K27M mutant sequence oligonucleotide and (b) non-tumor human CSF alone. (DOCX 268 kb)

Supplemental Information. Detailed methods and H3F3A K27M assay design. (DOCX 28 kb)

Supplemental Information. Detailed methods and H3F3A K27M assay design. (DOCX 28 kb)

Figure S1. Serial dilution of K27M mutant oligonucleotide in constant background of wild-type DNA demonstrates consistent detection down to at least 2% VAF under typical experimental conditions, with the possibility of detection at even lower VAF under ideal conditions. One such dilution series is shown above, with (a) showing number of droplets positive for mutant or wild-type H3F3A sequence and (b) showing the corresponding VAF values. Figure S2. Plot of droplets (blue â positive mutant H3F3A K27M, green â positive wildtype H3F3A, grey â negative droplets) from ddPCR performed on (a) non-tumor human CSF spiked with synthetic K27M mutant sequence oligonucleotide and (b) non-tumor human CSF alone. (DOCX 268 kb)