Medulloblastoma (MB) comprises a group of heterogeneous paediatric embryonal neoplasms of the hindbrain with strong links to early development of the hindbrain [1][2][3][4] . Mutations that activate Sonic hedgehog signalling lead to Sonic hedgehog MB in the upper rhombic lip (RL) granule cell lineage [5][6][7][8] . By contrast, mutations that activate WNT signalling lead to WNT MB in the lower RL 9,10 . However, little is known about the more commonly occurring group 4 (G4) MB, which is thought to arise in the unipolar brush cell lineage 3,4 . Here we demonstrate that somatic mutations that cause G4 MB converge on the core binding factor alpha (CBFA) complex and mutually exclusive alterations that affect CBFA2T2, CBFA2T3, PRDM6, UTX and OTX2. CBFA2T2 is expressed early in the progenitor cells of the cerebellar RL subventricular zone in Homo sapiens, and G4 MB transcriptionally resembles these progenitors but are stalled in developmental

Background: Polymerase chain reaction (PCR) has well established advantages over culture for diagnosis of herpes viruses, but its technical complexity has limited its widespread application. However, recent methodological advances have rendered PCR more applicable to routine practice. Aim: To compare automated PCR with viral culture for diagnosis of genital herpes. Methods: We studied 236 patients presenting with clinical features suggestive of genital herpes at an inner city genitourinary medicine clinic. Two swabs were taken from each patient. Cell culture and typing were performed by standard methods. Automated PCR was performed using the LightCycler instrument and the infecting viral type was determined by restriction endonuclease digestion of amplicons. Results: 109 patients (46%) had a positive test for herpes simplex virus (HSV). In 88, both PCR and culture were positive; in 21 PCR only was positive. With both detection methods, lesion duration and morphology were associated with HSV detection. Compared with culture alone, use of PCR increased sensitivity by 13.3% in specimens from vesicular lesions, by 27.4% from ulcerative lesions, and by 20.0% from crusting lesions. Conclusions: We advocate adoption of automated PCR as an efficient HSV detection and typing method for diagnosis of genital herpes in routine clinical practice. PCR allowed rapid laboratory confirmation of the diagnosis and increased the overall HSV detection rate by 24%.

Tumorigenic and non-neoplastic tissue injury occurs via the ischemic microenvironment defined by low oxygen, pH, and nutrients due to blood supply malfunction. Ischemic conditions exist within regions of pseudopalisading necrosis, a pathological hallmark of glioblastoma (GBM), the most common primary malignant brain tumor in adults. To recapitulate the physiologic microenvironment found in GBM tumors and tissue injury, we developed an in vitro ischemic model and identified chromodomain helicase DNA-binding protein 7 (CHD7) as a novel ischemia-regulated gene. Point mutations in the CHD7 gene are causal in CHARGE syndrome (a developmental disorder causing coloboma, heart defects, atresia choanae, retardation of growth, and genital and ear anomalies) and interrupt the epigenetic functions of CHD7 in regulating neural stem cell maintenance and development. Using our ischemic system, we observed microenvironment-mediated decreases in CHD7 expression in brain tumor-initiating cells and neural stem cells. Validating our approach, CHD7 was suppressed in the perinecrotic niche of GBM patient and xenograft sections, and an interrogation of patient gene expression datasets determined correlations of low CHD7 with increasing glioma grade and worse patient outcomes. Segregation of GBM by molecular subtype revealed a novel observation that CHD7 expression is elevated in proneural versus mesenchymal GBM. Genetic targeting of CHD7 and subsequent gene ontology analysis of RNA sequencing data indicated angiogenesis as a primary biological function affected by CHD7 expression changes. We validated this finding in tube-formation assays and vessel formation in orthotopic GBM models. Together, our data provide further understanding of molecular responses to ischemia and a novel function of CHD7 in regulating angiogenesis in both neoplastic and nonneoplastic systems. STEM CELLS 2019;37:453-462 SIGNIFICANCE STATEMENTThe epigenetic modifier chromodomain helicase DNA-binding protein 7 (CHD7) is known to have important roles in development, neural stem cell maintenance, and some cancers. The study finds for the first time that ischemic microenvironments repress CHD7 levels in neural progenitors and brain tumor-initiating cells/cancer stem cells. Results of the study also demonstrate that reduced levels of CHD7 in neural progenitors or brain tumor-initiating cells increase angiogenesis. The results may have important implications for development, stem cells, and cancer.

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Introduction: Pediatric high-grade gliomas (pHGGs) are routinely fatal with a median overall survival (OS) at recurrence of 5.6 months (mos). A safe, effective immunotherapy for pHGG has eluded investigators. Oncolytic HSV-1 G207 contains mutations that prevent a productive infection of normal cells but permit replication in tumor cells. In addition to direct tumor cell lysis, G207 activates innate/adaptive immune cells and promotes cross-presentation of tumor antigens to generate an anti-tumor immune response. A 5 Gy radiation dose increases viral replication and spread. We evaluated the safety and efficacy of G207 alone and combined with radiation in children with progressive supratentorial HGG (NCT02457845). Methods: We employed a 3 + 3 design with 4 cohorts. Children 3-18 years old with biopsy-confirmed HGG underwent stereotactic placement of up to four intratumoral catheters. The next day, we administered 107 or 108 plaque-forming units (pfu) of G207 by controlled rate infusion over 6 hours. Within 24 hours of G207, patients in dose level 3 and 4 received 5 Gy to the gross tumor volume. The primary objective was safety/tolerability. We assessed secondary objectives of virus shedding in blood, saliva and conjunctiva by PCR, response by MRI and evaluation of matched pre- and post-G207 tissue for tumor-infiltrating lymphocytes (TILs), and seroconversion by immunofluorescence assay. Results: 12 patients (age range 7-18) with progressive, IDH wild-type pHGG received G207. At screening, 10 patients had tumors with a bi-perpendicular sum ≥ 5.5 cm, 3 had multi-focal disease, 8 had failed ≥ 2 prior treatment regimens, and 4 had failed ≥ 3 regimens. 3-4 catheters (44 total) were placed safely throughout the cerebrum and resulted in no neurologic sequelae. G207 alone or with radiation was safe and tolerable in all patients with no dose-limiting toxicities, attributable grade 3 or 4 toxicities/serious adverse events, or evidence of virus shedding. 11 participants had radiographic, neuropathologic, and/or clinical responses. Median OS was 12.2 mos (95% CI 8.0, 16.4). Thus far, 36% of patients have lived >18 mos, the median OS for newly diagnosed pHGG. Compared to patients who seroconverted post-G207 (n=3), patients with baseline HSV-1 antibodies (n=3) had a shorter median survival: 5.1 mos (3.0, 7.2) vs 18.3 mos (9.2, 27.4). G207 significantly increased CD4+ and CD8+ TILs. Conclusions: G207 alone and combined with radiation was tolerable and safe with evidence of responses in children with pHGG. The promising median OS (12.2 mos) compares favorably with historical data (5.6 mos). Baseline HSV-1 seropositivity and seroconversion are potential biomarkers of treatment response that require further investigation. Importantly, G207 converted ‘cold' tumors to ‘hot' with a dramatic increase in TILs. A multi-institutional Phase II clinical trial of G207 in pHGG is forthcoming (NCT04482933). Citation Format: Gregory K. Friedman, James M. Johnston, Asim K. Bag, Joshua D. Bernstock, Rong Li, Inmaculada Aban, Kara Kachurak, Li Nan, Kyung-Don Kang, Stacie Totsch, Charles Schlappi, Allison M. Martin, Devang Pastakia, Sameer Farouk Sait, Yasmin Khakoo, Matthias A. Karajannis, Karina Woodling, Joshua D. Palmer, Diana S. Osorio, Jeffrey Leonard, Mohamed S. Abdelbaki, Avi Madan-Swain, T. Prescott Atkinson, Richard J. Whitley, John B. Fiveash, James M. Markert, G. Yancey Gillespie. Phase I immunovirotherapy trial of oncolytic HSV-1 G207 alone or combined with radiation in pediatric high-grade glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT018.