Glycolytic enzymes localize to ribonucleoprotein granules in Drosophila germ cells, bind Tudor and protect from transposable elements.

Germ cells give rise to all cell lineages in the next-generation and are responsible for the continuity of life. In a variety of organisms, germ cells and stem cells contain large ribonucleoprotein granules. Although these particles were discovered more than 100 years ago, their assembly and functions are not well understood. Here we report that glycolytic enzymes are components of these granules in Drosophila germ cells and both their mRNAs and the enzymes themselves are enriched in germ cells. We show that these enzymes are specifically required for germ cell development and that they protect their genomes from transposable elements, providing the first link between metabolism and transposon silencing. We further demonstrate that in the granules, glycolytic enzymes associate with the evolutionarily conserved Tudor protein. Our biochemical and single-particle EM structural analyses of purified Tudor show a flexible molecule and suggest a mechanism for the recruitment of glycolytic enzymes to the granules. Our data indicate that germ cells, similarly to stem cells and tumor cells, might prefer to produce energy through the glycolytic pathway, thus linking a particular metabolism to pluripotency.

Phase I Study of High-Dose L-methylfolate in Combination with Temozolomide and Bevacizumab in Recurrent IDH wild-type High-Grade Glioma.

Purpose: IDH mutations in low-grade gliomas (LGGs) results in improved survival and DNA hypermethylation compared to IDH wild-type LGGs. IDH-mutant LGGs become hypomethylated during progression. It's uncertain if methylation changes occur during IDH wild-type GBM progression and if the methylome can be reprogrammed. This phase I study evaluated the safety, tolerability, efficacy and methylome changes after L-methylfolate (LMF) treatment, in combination with temozolomide and bevacizumab in patients with recurrent high-grade glioma. Patients and Methods: Fourteen patients total, 13 with GBM, one with anaplastic astrocytoma, all IDH wild-type were enrolled in the study. All patients received LMF at either 15, 30, 60, or 90 mg daily plus temozolomide (75mg/m2 5 days per month) and bevacizumab (10mg/kg every two weeks). Results: No MTD was identified. LMF treated had mOS of 9.5 months (95% CI, 9.1-35.4) comparable to bevacizumab historical control 8.6 months (95% CI, 6.8-10.8). Six patients treated with LMF survived more than 650 days. Across all treatment doses the most adverse events were diarrhea (7%, 1 patient, grade 2), reflux (7%, 1 patient, grade 2), and dysgeusia (7%, 1 patient, grade 2). In the six brains donated at death, there was a 25% increase in DNA methylated CpGs compared to the paired initial tumor. Conclusions: LMF in combination with temozolomide and bevacizumab was well tolerated in patients with recurrent IDH wild-type high-grade glioma. This small study did not establish a superior efficacy with addition of LMF compared to standard bevacizumab therapy, however, this study did show methylome reprogramming in high-grade glioma.

Occurrence of bisphenol A in indoor dust from two locations in the eastern United States and implications for human exposures.

Bisphenol A (BPA) is used in the production of polycarbonate plastics and epoxy resins, which are used in many consumer products. Sources of human exposures to BPA include packaged and canned food products, indoor air, and dust ingestion. Information on the relative contributions of the pathways to BPA exposures is limited. In this study, we measured concentrations BPA in indoor dust collected from two locations in the Eastern United States and evaluated the contribution of dust to total BPA exposures. BPA was found in 95% of the dust samples analyzed (n = 56) at concentrations ranging from <0.5 to 10,200 ng/g (mean 843; median 422). The median values for BPA intake by way of the ingestion of dust by adults and toddlers were calculated to be 0.35 and 5.63 ng/kg body weight/day. These estimated exposure doses of BPA through dust ingestion are of the same order of magnitude as the recently reported low concentrations that induced health effects in laboratory animal studies. The contribution of dust to total human BPA intake was estimated to be <1%, however, suggesting that dietary intake is the predominant source of exposures in humans.

Novel role of specific Tudor domains in Tudor-Aubergine protein complex assembly and distribution during Drosophila oogenesis.

Germ cells give rise to the next generation and contain ribonucleoprotein particles, germ granules. In these granules, Piwi protein Aubergine has been shown to interact with Tudor protein in Drosophila. Tudor protein has 11 Tudor domains and it has been unclear to what extent all these domains are involved in the interaction with Aubergine. Here we present direct biochemical evidence that Tudor-Aubergine interaction surface is composed of different Tudor domains including those that have not been previously implicated in Aubergine recognition. Furthermore, we show that specific single Tudor domains determine localization of Tudor complex to different sites in ovarian germ cells. Our data suggest that multiple Tudor domains of germline proteins from various species are redundantly used for interaction with the same protein partner during germline development.

Targeting the IGF1R/PI3K/AKT Pathway Sensitizes Ewing Sarcoma to BET Bromodomain Inhibitors.

Inhibitors of the bromodomain and extra-terminal domain (BET) family proteins modulate EWS-FLI1 activities in Ewing sarcoma. However, the efficacy of BET inhibitors as a monotherapy was moderate and transient in preclinical models. The objective of this study was to identify the mechanisms mediating intrinsic resistance to BET inhibitors and develop more effective combination treatments for Ewing sarcoma. Using a panel of Ewing sarcoma cell lines and patient-derived xenograft lines (PDX), we demonstrated that IGF1R inhibitors synergistically increased sensitivities to BET inhibitors and induced potent apoptosis when combined with BET inhibitors. Constitutively activated AKT significantly protected Ewing sarcoma cells against BET inhibitors, suggesting that IGF1R regulates responsiveness to BET inhibitors mainly through the PI3K/AKT pathway. Although two Ewing sarcoma cell lines were resistant to IGF1R inhibitors, they retained synergistic response to a combination of BET inhibitors and mTOR inhibitors, suggesting that BET proteins, when IGF1R is not functional, cross-talk with its downstream molecules. Furthermore, the combination of a BET inhibitor and an IGF1R inhibitor induced potent and durable response in xenograft tumors, whereas either agent alone was less effective. Taken together, our results suggest that IGF1R and the downstream PI3K/AKT/mTOR kinase cascade mediate intrinsic resistance to BET inhibitors in Ewing sarcoma. These results provide the proof-of-concept for combining BET inhibitors with agents targeting the IGF1R pathway for treating advanced Ewing sarcoma.

The MAPK Pathway Regulates Intrinsic Resistance to BET Inhibitors in Colorectal Cancer.

Purpose: The bromodomain and extra-terminal domain (BET) family proteins are epigenetic readers for acetylated histone marks. Emerging BET bromodomain inhibitors have exhibited antineoplastic activities in a wide range of human cancers through suppression of oncogenic transcription factors, including MYC. However, the preclinical activities of BET inhibitors in advanced solid cancers are moderate at best. To improve BET-targeted therapy, we interrogated mechanisms mediating resistance to BET inhibitors in colorectal cancer.Experimental Design: Using a panel of molecularly defined colorectal cancer cell lines, we examined the impact of BET inhibition on cellular proliferation and survival as well as MYC activity. We further tested the ability of inhibitors targeting the RAF/MEK/ERK (MAPK) pathway to enhance MYC suppression and circumvent intrinsic resistance to BET inhibitors. Key findings were validated using genetic approaches.Results: BET inhibitors as monotherapy moderately reduced colorectal cancer cell proliferation and MYC expression. Blockade of the MAPK pathway synergistically sensitized colorectal cancer cells to BET inhibitors, leading to potent apoptosis and MYC downregulation in vitro and in vivo A combination of JQ1 and trametinib, but neither agent alone, induced significant regression of subcutaneous colorectal cancer xenografts.Conclusions: Our findings suggest that the MAPK pathway confers intrinsic resistance to BET inhibitors in colorectal cancer and propose an effective combination strategy for the treatment of colorectal cancer. Clin Cancer Res; 23(8); 2027-37. ©2016 AACR.

BET bromodomain inhibitors suppress EWS-FLI1-dependent transcription and the IGF1 autocrine mechanism in Ewing sarcoma.

Ewing sarcoma is driven by characteristic chromosomal translocations between the EWSR1 gene with genes encoding ETS family transcription factors (EWS-ETS), most commonly FLI1. However, direct pharmacological inhibition of transcription factors like EWS-FLI1 remains largely unsuccessful. Active gene transcription requires orchestrated actions of many epigenetic regulators, such as the bromodomain and extra-terminal domain (BET) family proteins. Emerging BET bromodomain inhibitors have exhibited promising antineoplastic activities via suppression of oncogenic transcription factors in various cancers. We reasoned that EWS-FLI1-mediated transcription activation might be susceptible to BET inhibition. In this study, we demonstrated that small molecule BET bromodomain inhibitors repressed EWS-FLI1-driven gene signatures and downregulated important target genes. However, expression of EWS-FLI1 was not significantly affected. Repression of autocrine IGF1 by BET inhibitors led to significant inhibition of the IGF1R/AKT pathway critical to Ewing sarcoma cell proliferation and survival. Consistently, BET inhibitors impaired viability and clonogenic survival of Ewing sarcoma cell lines and blocked EWS-FLI1-induced transformation of mouse NIH3T3 fibroblast cells. Selective depletion of individual BET genes partially phenocopied the actions of BET inhibitors. Finally, the prototypical BET inhibitor, JQ1, significantly repressed Ewing sarcoma xenograft tumor growth. These findings suggest therapeutic potential of BET inhibitors in Ewing sarcoma and highlight an emerging paradigm of using epigenetic agents to treat cancers driven by fusion transcription factors.

Devic's neuromyelitis optica: a clinicopathological review of the literature in connection with a case showing fatal dysautonomia.

A new clinicopathological case of Devic's neuromyelitis optica, including unusual predominant clinical features (fatal dysautonomia) is described herein: pathological examination showed extensive and severe demyelination mainly involving the optic nerves, the medulla, and spinal cord, which was particularly pronounced in the thoracic segments, and thus explained the unusual vegetative symptomatology. In a review of 45 clinicopathological cases described in the literature as Devic's disease (DD), it turned out that only 22 cases, including the patient described here, fulfilled the recently defined diagnostic criteria [Devic 1980]. Among the other 23 cases, 15 did not fulfill the criteria because of the occurrence of relapses, 3 others had 2 separate pathological locations within the spinal cord, and the remaining 5 showed evidence that a disease other than DD was involved. Among the 22 cases which were definitely taken to be DD, the mean age at clinical onset was 39 years (+/- 14); it was characterized by acute bilateral visual loss and transversal myelitis which gradually led either to death or to partial or complete recovery. The pathological lesions, which mostly were located along the optic tracts and the spinal cord, were characterized by demyelination with inflammation and necrosis; in some cases the pathological process diffused into the medulla (8/22 cases) or the whole brainstem (4/22 cases). The cavitation of the spinal cord was not a key feature of the diagnosis, since it was observed on only 9/22 cases. Devic's neuromyelitis optica may be a separate nosological entity which differs from multiple sclerosis.

[Benign papillomatosis of Wirsung's duct associated with adenomyoma of the distal common bile duct]. / Papillomatose bénigne du canal de Wirsung associée à un adénomyome du bas cholédoque.

We report the case of a 42 year-old man, with a history of alcohol and tobacco abuse, who was referred for suspicion of carcinoma of the head of pancreas. Pancreatoduodenectomy was performed. Histological study demonstrated obstructive benign adenomyoma of the distal common bile duct associated with villous papillomatosis of the pancreatic ducts and patchy lesions of chronic pancreatitis. Two years later, the patient was completely asymptomatic, but the risk of malignant transformation of villous papillomatosis requires periodic examination of the remaining pancreas and biliary tract.

[A case of papillary tumor of the pancreas]. / Un cas de tumeur papillaire du pancréas.

The authors report a case of papillary cystic tumor which is a rare form of pancreatic tumor of exocrine origin. A young woman presenting with a painful abdominal mass was operated on for a tumor of the head of the pancreas. Gross and microscopic examination revealed a papillary cystic tumor of the pancreas. Tests for histologic markers found progesterone receptors. A review of the literature found approximately 100 cases of papillary cystic tumors of the pancreas. This type of tumor is seen mainly in young women presenting with an abdominal mass. After complete resection of the tumor, these patients usually make an uneventful recovery. Since regrowth of the tumor may occur these patients should be submitted to prolonged follow-up.

[Interstitial pneumopathy revealing type B Niemann Pick disease in an adult]. / Pneumopathie interstitielle révélatrice d'une maladie de Niemann-Pick de type B chez un adulte.

INTRODUCTION: Niemann-Pick disease is an autosomal recessive disorder due to partial or total deficit in sphingomyelinase. EXEGESIS: We report a case of type B Niemann-Pick disease revealed by pneumonia and splenomegaly associated with blue histiocyte syndrome. Investigations that were done 2 years prior to diagnosis had shown the existence of isolated chronic lipid pneumonia which is specific of overloading. CONCLUSION: Though diagnosis is based on biochemistry, the morphology of alveolar histiocytes after simple bronchoalveolar lavage is of value.

[Characteristic ultrastructural features of parathyroid gland cancer. A report on 9 cases (author's transl)]. / Particularités ultrastructurales du cancer des glandes parathyroïdes. A propos de 9 observations.

Authors have studied 9 parathyroid carcinomas under electron microscope (3% of their parathyroid lesions). Two types of the ultrastructural features were significant: 1) architectural: basal membrane abnormalities, sparse desmosomes, absence of nerve fiber; 2) cytological: nuclei atypism and abnormal mitochondria in tumors cells resembling dark chief cells. Also noticed were signs of cell hyperactivity and/or trouble in parathormon production or regulation.

[Recurrent osteitis and Coxiella burnetii: the relation to chronic recurrent multifocal osteomyelitis]. / Ostéites récidivantes et Coxiella burnetti: relation avec l'ostéomyélite chronique récurrente multifocale.

UNLABELLED: Chronic recurrent multifocal osteomyelitis (CRMO) is a disorder of suspected--but unproved-infectious etiology. OBSERVATION: A girl presented with a typical CRMO involving successively the left fibula, radius, humerus and the right carpus. A Coxiella burnetii infection was indicated during the first attack. Two recurrences occurred in spite of suitable antibiotic treatment and with negative infectious investigation. Two months after stopping antibiotic treatment, a new recurrence associated with antibodies increase and positive bone culture occurred. CONCLUSION: Coxiella burnetii can initiate a CRMO. The mechanism involved is probably a delayed hypersensitivity. CRMO would therefore be the first type of reactive osteitis.

[Giant breast fibroadenoma]. / Le fibro-adénome mammaire géant.

We report a case of giant fibroadenoma of the breast, "juvenile type", in a 15 year old girl. We study the characteristics of this tumor that occurs in adolescents females, the differential diagnosis with cystosarcoma phylloide and benign virginal hypertrophy, and treatment. Follow-up is recommended because of possible recurrent tumor of a short time.

[Infantile polycystic hepatorenal disease. 2 consecutive cases in a patient. Ultrasonic diagnosis and indication for elective pregnancy termination]. / Maladie polykystique hépatorénale infantile. Deux cas consécutifs chez une patiente. Diagnostic échographique et indication d'interruption médicale de grossesse.

We report a case of infantile polycystic kidney disease, during the course of two consecutive pregnancies in the same woman. Observed rates of recurrence in families at risk is higher than theoretical rates (25%). Antenatal ultrasound can show signs of bilateral involvement, which is always lethal and generally leads to elective termination of pregnancy. Diagnosis can rarely be made before 24 weeks of pregnancy.

[Visceral leishmaniasis associated with Hodgkin's disease: diagnostic difficulties]. / Leishmaniose viscérale associée à une maladie de Hodgkin: difficultés diagnostiques.

A case of Hodgkin's disease associated from the start with visceral leishmaniasis in the absence of antitumoral treatment shows that leishmaniasis is a severe opportunistic infection in endemic areas and can be masked by the tumoral syndrome of an underlying pathology. Conversely, patients with visceral leishmaniasis must be investigated for a cause of immunosuppression with, in particular, biopsy of accessible lymph nodes. The exceptionally favourable course of this particular case deserved to be high-lighted.

An in vivo crosslinking approach to isolate protein complexes from Drosophila embryos.

Many cellular processes are controlled by multisubunit protein complexes. Frequently these complexes form transiently and require native environment to assemble. Therefore, to identify these functional protein complexes, it is important to stabilize them in vivo before cell lysis and subsequent purification. Here we describe a method used to isolate large bona fide protein complexes from Drosophila embryos. This method is based on embryo permeabilization and stabilization of the complexes inside the embryos by in vivo crosslinking using a low concentration of formaldehyde, which can easily cross the cell membrane. Subsequently, the protein complex of interest is immunopurified followed by gel purification and analyzed by mass spectrometry. We illustrate this method using purification of a Tudor protein complex, which is essential for germline development. Tudor is a large protein, which contains multiple Tudor domains--small modules that interact with methylated arginines or lysines of target proteins. This method can be adapted for isolation of native protein complexes from different organisms and tissues.