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Sample records for damage checkpoint stimulation

  1. Role of the Checkpoint Clamp in DNA Damage Response

    Directory of Open Access Journals (Sweden)

    Mihoko Kai

    2013-01-01

    Full Text Available DNA damage occurs during DNA replication, spontaneous chemical reactions, and assaults by external or metabolism-derived agents. Therefore, all living cells must constantly contend with DNA damage. Cells protect themselves from these genotoxic stresses by activating the DNA damage checkpoint and DNA repair pathways. Coordination of these pathways requires tight regulation in order to prevent genomic instability. The checkpoint clamp complex consists of Rad9, Rad1 and Hus1 proteins, and is often called the 9-1-1 complex. This PCNA (proliferating cell nuclear antigen-like donut-shaped protein complex is a checkpoint sensor protein that is recruited to DNA damage sites during the early stage of the response, and is required for checkpoint activation. As PCNA is required for multiple pathways of DNA metabolism, the checkpoint clamp has also been implicated in direct roles in DNA repair, as well as in coordination of the pathways. Here we discuss roles of the checkpoint clamp in DNA damage response (DDR.

  2. Orchestration of DNA Damage Checkpoint Dynamics across the Human Cell Cycle.

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    Chao, Hui Xiao; Poovey, Cere E; Privette, Ashley A; Grant, Gavin D; Chao, Hui Yan; Cook, Jeanette G; Purvis, Jeremy E

    2017-11-22

    Although molecular mechanisms that prompt cell-cycle arrest in response to DNA damage have been elucidated, the systems-level properties of DNA damage checkpoints are not understood. Here, using time-lapse microscopy and simulations that model the cell cycle as a series of Poisson processes, we characterize DNA damage checkpoints in individual, asynchronously proliferating cells. We demonstrate that, within early G1 and G2, checkpoints are stringent: DNA damage triggers an abrupt, all-or-none cell-cycle arrest. The duration of this arrest correlates with the severity of DNA damage. After the cell passes commitment points within G1 and G2, checkpoint stringency is relaxed. By contrast, all of S phase is comparatively insensitive to DNA damage. This checkpoint is graded: instead of halting the cell cycle, increasing DNA damage leads to slower S phase progression. In sum, we show that a cell's response to DNA damage depends on its exact cell-cycle position and that checkpoints are phase-dependent, stringent or relaxed, and graded or all-or-none. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Top3 processes recombination intermediates and modulates checkpoint activity after DNA damage

    DEFF Research Database (Denmark)

    Mankouri, Hocine W; Hickson, Ian D

    2006-01-01

    Mutation of TOP3 in Saccharomyces cerevisiae causes poor growth, hyperrecombination, and a failure to fully activate DNA damage checkpoints in S phase. Here, we report that overexpression of a dominant-negative allele of TOP3, TOP3(Y356F), which lacks the catalytic (decatenation) activity of Top3......, the catalytic activity of Top3 is not required for DNA damage checkpoint activation, but it is required for normal S-phase progression after DNA damage. We also present evidence that the checkpoint-mediated cell cycle delay and persistence of X-shaped DNA molecules resulting from overexpression of TOP3(Y356F......) are downstream of Rad51 function. We propose that Top3 functions in S phase to both process homologous recombination intermediates and modulate checkpoint activity....

  4. rad-Dependent response of the chk1-encoded protein kinase at the DNA damage checkpoint

    NARCIS (Netherlands)

    Walworth, N.C.; Bernards, R.A.

    1996-01-01

    Exposure of eukaryotic cells to agents that generate DNA damage results in transient arrest of progression through the cell cycle. In fission yeast, the DNA damage checkpoint associated with cell cycle arrest before mitosis requires the protein kinase p56chk1. DNA damage induced by ultraviolet

  5. NEK11: linking CHK1 and CDC25A in DNA damage checkpoint signaling

    DEFF Research Database (Denmark)

    Sørensen, Claus Storgaard; Melixetian, Marina; Klein, Ditte Kjaersgaard

    2010-01-01

    The DNA damage induced G(2)/M checkpoint is an important guardian of the genome that prevents cell division when DNA lesions are present. The checkpoint prevents cells from entering mitosis by degrading CDC25A, a key CDK activator. CDC25A proteolysis is controlled by direct phosphorylation events...... is required for beta-TrCP mediated CDC25A polyubiquitylation and degradation. The activity of NEK11 is in turn controlled by CHK1 that activates NEK11 via phosphorylation on serine 273. Since inhibition of NEK11 activity forces checkpoint-arrested cells into mitosis and cell death, NEK11 is, like CHK1...

  6. The anaphase inhibitor of Saccharomyces cerevisiae Pds1p is a target of the DNA damage checkpoint pathway

    International Nuclear Information System (INIS)

    Cohen-Fix, O.; Koshland, D.

    1997-01-01

    Inhibition of DNA replication and physical DNA damage induce checkpoint responses that arrest cell cycle progression at two different stages. In Saccharomyces cerevisiae, the execution of both checkpoint responses requires the Mec1 and Rad53 proteins. This observation led to the suggestion that these checkpoint responses are mediated through a common signal transduction pathway. However, because the checkpoint-induced arrests occur at different cell cycle stages, the downstream effectors mediating these arrests are likely to be distinct. We have previously shown that the S. cerevisiae protein Pds1p is an anaphase inhibitor and is essential for cell cycle arrest in mitosis in the presence DNA damage. Herein we show that DNA damage, but not inhibition of DNA replication, induces the phosphorylation of Pds1p. Analyses of Pds1p phosphorylation in different checkpoint mutants reveal that in the presence of DNA damage, Pds1p is phosphorylated in a Mec1p- and Rad9p-dependent hut Rad53p-independent manner. Our data place Pds1p and Rad53p on parallel branches of the DNA damage checkpoint pathway. We suggest that Pds1p is a downstream target of the DNA damage checkpoint pathway and that it is involved in implementing the DNA damage checkpoint arrest specifically in mitosis

  7. [Sea urchin embryo, DNA-damaged cell cycle checkpoint and the mechanisms initiating cancer development].

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    Bellé, Robert; Le Bouffant, Ronan; Morales, Julia; Cosson, Bertrand; Cormier, Patrick; Mulner-Lorillon, Odile

    2007-01-01

    Cell division is an essential process for heredity, maintenance and evolution of the whole living kingdom. Sea urchin early development represents an excellent experimental model for the analysis of cell cycle checkpoint mechanisms since embryonic cells contain a functional DNA-damage checkpoint and since the whole sea urchin genome is sequenced. The DNA-damaged checkpoint is responsible for an arrest in the cell cycle when DNA is damaged or incorrectly replicated, for activation of the DNA repair mechanism, and for commitment to cell death by apoptosis in the case of failure to repair. New insights in cancer biology lead to two fundamental concepts about the very first origin of cancerogenesis. Cancers result from dysfunction of DNA-damaged checkpoints and cancers appear as a result of normal stem cell (NCS) transformation into a cancer stem cell (CSC). The second aspect suggests a new definition of "cancer", since CSC can be detected well before any clinical evidence. Since early development starts from the zygote, which is a primary stem cell, sea urchin early development allows analysis of the early steps of the cancerization process. Although sea urchins do not develop cancers, the model is alternative and complementary to stem cells which are not easy to isolate, do not divide in a short time and do not divide synchronously. In the field of toxicology and incidence on human health, the sea urchin experimental model allows assessment of cancer risk from single or combined molecules long before any epidemiologic evidence is available. Sea urchin embryos were used to test the worldwide used pesticide Roundup that contains glyphosate as the active herbicide agent; it was shown to activate the DNA-damage checkpoint of the first cell cycle of development. The model therefore allows considerable increase in risk evaluation of new products in the field of cancer and offers a tool for the discovery of molecular markers for early diagnostic in cancer biology

  8. The amount of DNA damage needed to activate the radiation-induced G2 checkpoint varies between single cells

    International Nuclear Information System (INIS)

    Tkacz-Stachowska, Kinga; Lund-Andersen, Christin; Velissarou, Angeliki; Myklebust, June H.; Stokke, Trond; Syljuåsen, Randi G.

    2011-01-01

    Background and purpose: The radiation-induced G2 checkpoint helps facilitate DNA repair before cell division. However, recent work has revealed that human cells often escape the G2 checkpoint with unrepaired DNA breaks. The purpose was to explore whether G2 checkpoint activation occurs according to a threshold level of DNA damage. Materials and methods: G2 checkpoint activation was assayed at 75–90 min and 24–48 h after X-ray irradiation of BJ diploid fibroblasts and U2OS osteosarcoma cells. Multiparameter flow cytometry with pacific blue barcoding, and flow cytometry-based sorting of phospho-H3 positive cells to microscope slides, were used to examine the DNA damage marker γ-H2AX in individual mitotic cells that had escaped the G2 checkpoint. Results: For all radiation doses and times tested, the number of γ-H2AX foci varied between individual mitotic cells. At 75 min the median levels of γ-H2AX in mitotic cells increased with higher radiation doses. At 24–48 h, following a prolonged G2 checkpoint, cells were more resistant to checkpoint re-activation by a second dose of radiation. Conclusion: Our results suggest that different amounts of DNA damage are needed to activate the G2 checkpoint in individual cells. Such single cell variation in checkpoint activation may potentially contribute to radiation-induced genomic instability.

  9. Biological significance of the focus on DNA damage checkpoint factors remained after irradiation of ionizing radiation

    International Nuclear Information System (INIS)

    Yamauchi, Motohiro; Suzuki, Keiji

    2005-01-01

    This paper reviews recent reports on the focus formation and participation to checkpoint of (such phosphorylated (P-d) as below) ATM and H2AX, MDC1, 53BP1 and NBS1, and discusses their role in DNA damage checkpoint induction mainly around authors' studies. When the cell is irradiated by ionizing radiation, the subtype histone like H2AX is P-d and the formed focus', seen in the nucleus on immuno-fluorographic observation, represents the P-d H2AX at the damaged site of DNA. The role of P-d ATM (the product of causative gene of ataxia-telangiectasia mutation, a protein kinase) has been first shown by laser beam irradiation. Described are discussions on the roles and functions after irradiation in focus formation and DNA damage checkpoint of P-d H2AX (a specific histone product by the radiation like γ-ray as above), P-d ATM, MDC1 (a mediator of DNA damage check point protein 1), 53BP1, (a p53 binding protein) and NBS1 (the product of the causative gene of Nijmegen Breakage Syndrome). Authors have come to point out the remained focal size increase as implications of the efficient repair of damaged DNA, and the second cycled p53 accumulation, of tumor suppression. Thus evaluation of biological significance of these aspects, scarcely noted hitherto, is concluded important. (S.I.)

  10. Oncogene-induced senescence is part of the tumorigenesis barrier imposed by DNA damage checkpoints

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    Bartkova, Jirina; Rezaei, Nousin; Liontos, Michalis

    2006-01-01

    Recent studies have indicated the existence of tumorigenesis barriers that slow or inhibit the progression of preneoplastic lesions to neoplasia. One such barrier involves DNA replication stress, which leads to activation of the DNA damage checkpoint and thereby to apoptosis or cell cycle arrest...... and senescence markers cosegregate closely. Thus, senescence in human preneoplastic lesions is a manifestation of oncogene-induced DNA replication stress and, together with apoptosis, provides a barrier to malignant progression....

  11. Loss of p53 induces M-phase retardation following G2 DNA damage checkpoint abrogation.

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    Minemoto, Yuzuru; Uchida, Sanae; Ohtsubo, Motoaki; Shimura, Mari; Sasagawa, Toshiyuki; Hirata, Masato; Nakagama, Hitoshi; Ishizaka, Yukihito; Yamashita, Katsumi

    2003-04-01

    Most cell lines that lack functional p53 protein are arrested in the G2 phase of the cell cycle due to DNA damage. When the G2 checkpoint is abrogated, these cells are forced into mitotic catastrophe. A549 lung adenocarcinoma cells, in which p53 was eliminated with the HPV16 E6 gene, exhibited efficient arrest in the G2 phase when treated with adriamycin. Administration of caffeine to G2-arrested cells induced a drastic change in cell phenotype, the nature of which depended on the status of p53. Flow cytometric and microscopic observations revealed that cells that either contained or lacked p53 resumed their cell cycles and entered mitosis upon caffeine treatment. However, transit to the M phase was slower in p53-negative cells than in p53-positive cells. Consistent with these observations, CDK1 activity was maintained at high levels, along with stable cyclin B1, in p53-negative cells. The addition of butyrolactone I, which is an inhibitor of CDK1 and CDK2, to the p53-negative cells reduced the floating round cell population and induced the disappearance of cyclin B1. These results suggest a relationship between the p53 pathway and the ubiquitin-mediated degradation of mitotic cyclins and possible cross-talk between the G2-DNA damage checkpoint and the mitotic checkpoint.

  12. Targeting neddylation induces DNA damage and checkpoint activation and sensitizes chronic lymphocytic leukemia B cells to alkylating agents.

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    Paiva, C; Godbersen, J C; Berger, A; Brown, J R; Danilov, A V

    2015-07-09

    Microenvironment-mediated upregulation of the B-cell receptor (BCR) and nuclear factor-κB (NF-κB) signaling in CLL cells resident in the lymph node and bone marrow promotes apoptosis evasion and clonal expansion. We recently reported that MLN4924 (pevonedistat), an investigational agent that inhibits the NEDD8-activating enzyme (NAE), abrogates stromal-mediated NF-κB pathway activity and CLL cell survival. However, the NAE pathway also assists degradation of multiple other substrates. MLN4924 has been shown to induce DNA damage and cell cycle arrest, but the importance of this mechanism in primary neoplastic B cells has not been studied. Here we mimicked the lymph node microenvironment using CD40 ligand (CD40L)-expressing stroma and interleukin-21 (IL-21) to find that inducing proliferation of the primary CLL cells conferred enhanced sensitivity to NAE inhibition. Treatment of the CD40-stimulated CLL cells with MLN4924 resulted in deregulation of Cdt1, a DNA replication licensing factor, and cell cycle inhibitors p21 and p27. This led to DNA damage, checkpoint activation and G2 arrest. Alkylating agents bendamustine and chlorambucil enhanced MLN4924-mediated DNA damage and apoptosis. These events were more prominent in cells stimulated with IL-21 compared with CD40L alone, indicating that, following NAE inhibition, the culture conditions were able to direct CLL cell fate from an NF-κB inhibition to a Cdt1 induction program. Our data provide insight into the biological consequences of targeting NAE in CLL and serves as further rationale for studying the clinical activity of MLN4924 in CLL, particularly in combination with alkylating agents.

  13. Comparison of checkpoint responses triggered by DNA polymerase inhibition versus DNA damaging agents

    International Nuclear Information System (INIS)

    Liu, J.-S.; Kuo, S.-R.; Melendy, Thomas

    2003-01-01

    To better understand the different cellular responses to replication fork pausing versus blockage, early DNA damage response markers were compared after treatment of cultured mammalian cells with agents that either inhibit DNA polymerase activity (hydroxyurea (HU) or aphidicolin) or selectively induce S-phase DNA damage responses (the DNA alkylating agents, methyl methanesulfonate (MMS) and adozelesin). These agents were compared for their relative abilities to induce phosphorylation of Chk1, H2AX, and replication protein A (RPA), and intra-nuclear focalization of γ-H2AX and RPA. Treatment by aphidicolin and HU resulted in phosphorylation of Chk1, while HU, but not aphidicolin, induced focalization of γ-H2AX and RPA. Surprisingly, pre-treatment with aphidicolin to stop replication fork progression, did not abrogate HU-induced γ-H2AX and RPA focalization. This suggests that HU may act on the replication fork machinery directly, such that fork progression is not required to trigger these responses. The DNA-damaging fork-blocking agents, adozelesin and MMS, both induced phosphorylation and focalization of H2AX and RPA. Unlike adozelesin and HU, the pattern of MMS-induced RPA focalization did not match the BUdR incorporation pattern and was not blocked by aphidicolin, suggesting that MMS-induced damage is not replication fork-dependent. In support of this, MMS was the only reagent used that did not induce phosphorylation of Chk1. These results indicate that induction of DNA damage checkpoint responses due to adozelesin is both replication fork and fork progression dependent, induction by HU is replication fork dependent but progression independent, while induction by MMS is independent of both replication forks and fork progression

  14. Comparison of checkpoint responses triggered by DNA polymerase inhibition versus DNA damaging agents

    Energy Technology Data Exchange (ETDEWEB)

    Liu, J.-S.; Kuo, S.-R.; Melendy, Thomas

    2003-11-27

    To better understand the different cellular responses to replication fork pausing versus blockage, early DNA damage response markers were compared after treatment of cultured mammalian cells with agents that either inhibit DNA polymerase activity (hydroxyurea (HU) or aphidicolin) or selectively induce S-phase DNA damage responses (the DNA alkylating agents, methyl methanesulfonate (MMS) and adozelesin). These agents were compared for their relative abilities to induce phosphorylation of Chk1, H2AX, and replication protein A (RPA), and intra-nuclear focalization of {gamma}-H2AX and RPA. Treatment by aphidicolin and HU resulted in phosphorylation of Chk1, while HU, but not aphidicolin, induced focalization of {gamma}-H2AX and RPA. Surprisingly, pre-treatment with aphidicolin to stop replication fork progression, did not abrogate HU-induced {gamma}-H2AX and RPA focalization. This suggests that HU may act on the replication fork machinery directly, such that fork progression is not required to trigger these responses. The DNA-damaging fork-blocking agents, adozelesin and MMS, both induced phosphorylation and focalization of H2AX and RPA. Unlike adozelesin and HU, the pattern of MMS-induced RPA focalization did not match the BUdR incorporation pattern and was not blocked by aphidicolin, suggesting that MMS-induced damage is not replication fork-dependent. In support of this, MMS was the only reagent used that did not induce phosphorylation of Chk1. These results indicate that induction of DNA damage checkpoint responses due to adozelesin is both replication fork and fork progression dependent, induction by HU is replication fork dependent but progression independent, while induction by MMS is independent of both replication forks and fork progression.

  15. DNA damage response and spindle assembly checkpoint function throughout the cell cycle to ensure genomic integrity.

    Directory of Open Access Journals (Sweden)

    Katherine S Lawrence

    2015-04-01

    Full Text Available Errors in replication or segregation lead to DNA damage, mutations, and aneuploidies. Consequently, cells monitor these events and delay progression through the cell cycle so repair precedes division. The DNA damage response (DDR, which monitors DNA integrity, and the spindle assembly checkpoint (SAC, which responds to defects in spindle attachment/tension during metaphase of mitosis and meiosis, are critical for preventing genome instability. Here we show that the DDR and SAC function together throughout the cell cycle to ensure genome integrity in C. elegans germ cells. Metaphase defects result in enrichment of SAC and DDR components to chromatin, and both SAC and DDR are required for metaphase delays. During persistent metaphase arrest following establishment of bi-oriented chromosomes, stability of the metaphase plate is compromised in the absence of DDR kinases ATR or CHK1 or SAC components, MAD1/MAD2, suggesting SAC functions in metaphase beyond its interactions with APC activator CDC20. In response to DNA damage, MAD2 and the histone variant CENPA become enriched at the nuclear periphery in a DDR-dependent manner. Further, depletion of either MAD1 or CENPA results in loss of peripherally associated damaged DNA. In contrast to a SAC-insensitive CDC20 mutant, germ cells deficient for SAC or CENPA cannot efficiently repair DNA damage, suggesting that SAC mediates DNA repair through CENPA interactions with the nuclear periphery. We also show that replication perturbations result in relocalization of MAD1/MAD2 in human cells, suggesting that the role of SAC in DNA repair is conserved.

  16. DNA damage checkpoint kinase ATM regulates germination and maintains genome stability in seeds.

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    Waterworth, Wanda M; Footitt, Steven; Bray, Clifford M; Finch-Savage, William E; West, Christopher E

    2016-08-23

    Genome integrity is crucial for cellular survival and the faithful transmission of genetic information. The eukaryotic cellular response to DNA damage is orchestrated by the DNA damage checkpoint kinases ATAXIA TELANGIECTASIA MUTATED (ATM) and ATM AND RAD3-RELATED (ATR). Here we identify important physiological roles for these sensor kinases in control of seed germination. We demonstrate that double-strand breaks (DSBs) are rate-limiting for germination. We identify that desiccation tolerant seeds exhibit a striking transcriptional DSB damage response during germination, indicative of high levels of genotoxic stress, which is induced following maturation drying and quiescence. Mutant atr and atm seeds are highly resistant to aging, establishing ATM and ATR as determinants of seed viability. In response to aging, ATM delays germination, whereas atm mutant seeds germinate with extensive chromosomal abnormalities. This identifies ATM as a major factor that controls germination in aged seeds, integrating progression through germination with surveillance of genome integrity. Mechanistically, ATM functions through control of DNA replication in imbibing seeds. ATM signaling is mediated by transcriptional control of the cell cycle inhibitor SIAMESE-RELATED 5, an essential factor required for the aging-induced delay to germination. In the soil seed bank, seeds exhibit increased transcript levels of ATM and ATR, with changes in dormancy and germination potential modulated by environmental signals, including temperature and soil moisture. Collectively, our findings reveal physiological functions for these sensor kinases in linking genome integrity to germination, thereby influencing seed quality, crucial for plant survival in the natural environment and sustainable crop production.

  17. Direct and indirect control of the initiation of meiotic recombination by DNA damage checkpoint mechanisms in budding yeast.

    Directory of Open Access Journals (Sweden)

    Bilge Argunhan

    Full Text Available Meiotic recombination plays an essential role in the proper segregation of chromosomes at meiosis I in many sexually reproducing organisms. Meiotic recombination is initiated by the scheduled formation of genome-wide DNA double-strand breaks (DSBs. The timing of DSB formation is strictly controlled because unscheduled DSB formation is detrimental to genome integrity. Here, we investigated the role of DNA damage checkpoint mechanisms in the control of meiotic DSB formation using budding yeast. By using recombination defective mutants in which meiotic DSBs are not repaired, the effect of DNA damage checkpoint mutations on DSB formation was evaluated. The Tel1 (ATM pathway mainly responds to unresected DSB ends, thus the sae2 mutant background in which DSB ends remain intact was employed. On the other hand, the Mec1 (ATR pathway is primarily used when DSB ends are resected, thus the rad51 dmc1 double mutant background was employed in which highly resected DSBs accumulate. In order to separate the effect caused by unscheduled cell cycle progression, which is often associated with DNA damage checkpoint defects, we also employed the ndt80 mutation which permanently arrests the meiotic cell cycle at prophase I. In the absence of Tel1, DSB formation was reduced in larger chromosomes (IV, VII, II and XI whereas no significant reduction was found in smaller chromosomes (III and VI. On the other hand, the absence of Rad17 (a critical component of the ATR pathway lead to an increase in DSB formation (chromosomes VII and II were tested. We propose that, within prophase I, the Tel1 pathway facilitates DSB formation, especially in bigger chromosomes, while the Mec1 pathway negatively regulates DSB formation. We also identified prophase I exit, which is under the control of the DNA damage checkpoint machinery, to be a critical event associated with down-regulating meiotic DSB formation.

  18. The effect of tributyltin chloride on Caenorhabditis elegans germline is mediated by a conserved DNA damage checkpoint pathway.

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    Cheng, Zhe; Tian, Huimin; Chu, Hongran; Wu, Jianjian; Li, Yingying; Wang, Yanhai

    2014-03-21

    Tributyltin (TBT), one of the environmental pollutants, has been shown to impact the reproduction of animals. However, due to the lack of appropriate animal model, analysis of the affected molecular pathways in germ cells is lagging and has been particularly challenging. In the present study, we investigated the effects of tributyltin chloride (TBTCL) on the nematode Caenorhabditis elegans germline. We show that exposure of C. elegans to TBTCL causes significantly elevated level of sterility and embryonic lethality. TBTCL exposure results in an increased number of meiotic DNA double-strand breaks in germ cells, subsequently leading to activated DNA damage checkpoint. Exposing C. elegans to TBTCL causes dose- and time-dependent germline apoptosis. This apoptotic response was blocked in loss-of-function mutants of hus-1 (op241), mrt-2 (e2663) and p53/cep-1 (gk138), indicating that checkpoints and p53 are essential for mediating TBTCL-induced germ cell apoptosis. Moreover, TBTCL exposure can inhibit germ cell proliferation, which is also mediated by the conserved checkpoint pathway. We thereby propose that TBT exhibits its effects on the germline by inducing DNA damage and impaired maintenance of genomic integrity. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  19. Silencing of poly(ADP-ribose) glycohydrolase sensitizes lung cancer cells to radiation through the abrogation of DNA damage checkpoint

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    Nakadate, Yusuke [Shien-Lab, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Department of Bioengineering, Graduate School of Engineering, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); Kodera, Yasuo; Kitamura, Yuka [Shien-Lab, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Tachibana, Taro [Department of Bioengineering, Graduate School of Engineering, Osaka City University, 3-3-138 Sugimoto, Sumiyoshi-ku, Osaka 558-8585 (Japan); Tamura, Tomohide [Division of Thoracic Oncology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan); Koizumi, Fumiaki, E-mail: fkoizumi@ncc.go.jp [Division of Thoracic Oncology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045 (Japan)

    2013-11-29

    Highlights: •Radiosensitization by PARG silencing was observed in multiple lung cancer cells. •PAR accumulation was enhanced by PARG silencing after DNA damage. •Radiation-induced G2/M arrest and checkpoint activation were impaired by PARG siRNA. -- Abstract: Poly(ADP-ribose) glycohydrolase (PARG) is a major enzyme that plays a role in the degradation of poly(ADP-ribose) (PAR). PARG deficiency reportedly sensitizes cells to the effects of radiation. In lung cancer, however, it has not been fully elucidated. Here, we investigated whether PARG siRNA contributes to an increased radiosensitivity using 8 lung cancer cell lines. Among them, the silencing of PARG induced a radiosensitizing effect in 5 cell lines. Radiation-induced G2/M arrest was largely suppressed by PARG siRNA in PC-14 and A427 cells, which exhibited significantly enhanced radiosensitivity in response to PARG knockdown. On the other hand, a similar effect was not observed in H520 cells, which did not exhibit a radiosensitizing effect. Consistent with a cell cycle analysis, radiation-induced checkpoint signals were not well activated in the PC-14 and A427 cells when treated with PARG siRNA. These results suggest that the increased sensitivity to radiation induced by PARG knockdown occurs through the abrogation of radiation-induced G2/M arrest and checkpoint activation in lung cancer cells. Our findings indicate that PARG could be a potential target for lung cancer treatments when used in combination with radiotherapy.

  20. NDR1 modulates the UV-induced DNA-damage checkpoint and nucleotide excision repair

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jeong-Min; Choi, Ji Ye [Department of Biological Science, Dong-A University, Busan (Korea, Republic of); Yi, Joo Mi [Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan (Korea, Republic of); Chung, Jin Woong; Leem, Sun-Hee; Koh, Sang Seok [Department of Biological Science, Dong-A University, Busan (Korea, Republic of); Kang, Tae-Hong, E-mail: thkang@dau.ac.kr [Department of Biological Science, Dong-A University, Busan (Korea, Republic of)

    2015-06-05

    Nucleotide excision repair (NER) is the sole mechanism of UV-induced DNA lesion repair in mammals. A single round of NER requires multiple components including seven core NER factors, xeroderma pigmentosum A–G (XPA–XPG), and many auxiliary effector proteins including ATR serine/threonine kinase. The XPA protein helps to verify DNA damage and thus plays a rate-limiting role in NER. Hence, the regulation of XPA is important for the entire NER kinetic. We found that NDR1, a novel XPA-interacting protein, modulates NER by modulating the UV-induced DNA-damage checkpoint. In quiescent cells, NDR1 localized mainly in the cytoplasm. After UV irradiation, NDR1 accumulated in the nucleus. The siRNA knockdown of NDR1 delayed the repair of UV-induced cyclobutane pyrimidine dimers in both normal cells and cancer cells. It did not, however, alter the expression levels or the chromatin association levels of the core NER factors following UV irradiation. Instead, the NDR1-depleted cells displayed reduced activity of ATR for some set of its substrates including CHK1 and p53, suggesting that NDR1 modulates NER indirectly via the ATR pathway. - Highlights: • NDR1 is a novel XPA-interacting protein. • NDR1 accumulates in the nucleus in response to UV irradiation. • NDR1 modulates NER (nucleotide excision repair) by modulating the UV-induced DNA-damage checkpoint response.

  1. The Interaction between Checkpoint Kinase 1 (Chk1) and the Minichromosome Maintenance (MCM) Complex Is Required for DNA Damage-induced Chk1 Phosphorylation*

    Science.gov (United States)

    Han, Xiangzi; Aslanian, Aaron; Fu, Kang; Tsuji, Toshiya; Zhang, Youwei

    2014-01-01

    Chk1 is an essential mediator of the DNA damage response and cell cycle checkpoint. However, how exactly Chk1 transduces the checkpoint signaling is not fully understood. Here we report the identification of the heterohexamic minichromosome maintenance (MCM) complex that interacts with Chk1 by mass spectrometry. The interaction between Chk1 and the MCM complex was reduced by DNA damage treatment. We show that the MCM complex, at least partially, contributes to the chromatin association of Chk1, allowing for immediate phosphorylation of Chk1 by ataxia telangiectasia mutated and Rad3-related (ATR) in the presence of DNA damage. Further, phosphorylation of Chk1 at ATR sites reduces the interaction between Chk1 and the MCM complex, facilitating chromatin release of phosphorylated Chk1, a critical step in the initiation and amplification of cell cycle checkpoint. Together, these data provide novel insights into the activation of Chk1 in response to DNA damage. PMID:25049228

  2. Roles of nibrin and ATM/ATR kinases on the G2 checkpoint under endogenous or radio-induced DNA damage

    Directory of Open Access Journals (Sweden)

    Katherine Marcelain

    2005-01-01

    Full Text Available Checkpoint response to DNA damage involves the activation of DNA repair and G2 lengthening subpathways. The roles of nibrin (NBS1 and the ATM/ATR kinases in the G2 DNA damage checkpoint, evoked by endogenous and radio-induced DNA damage, were analyzed in control, A-T and NBS lymphoblast cell lines. Short-term responses to G2 treatments were evaluated by recording changes in the yield of chromosomal aberrations in the ensuing mitosis, due to G2 checkpoint adaptation, and also in the duration of G2 itself. The role of ATM/ATR in the G2 checkpoint pathway repairing chromosomal aberrations was unveiled by caffeine inhibition of both kinases in G2. In the control cell lines, nibrin and ATM cooperated to provide optimum G2 repair for endogenous DNA damage. In the A-T cells, ATR kinase substituted successfully for ATM, even though no G2 lengthening occurred. X-ray irradiation (0.4 Gy in G2 increased chromosomal aberrations and lengthened G2, in both mutant and control cells. However, the repair of radio-induced DNA damage took place only in the controls. It was associated with nibrin-ATM interaction, and ATR did not substitute for ATM. The absence of nibrin prevented the repair of both endogenous and radio-induced DNA damage in the NBS cells and partially affected the induction of G2 lengthening.

  3. Upregulated ATM gene expression and activated DNA crosslink-induced damage response checkpoint in Fanconi anemia: implications for carcinogenesis.

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    Yamamoto, Kazuhiko; Nihrane, Abdallah; Aglipay, Jason; Sironi, Juan; Arkin, Steven; Lipton, Jeffrey M; Ouchi, Toru; Liu, Johnson M

    2008-01-01

    Fanconi anemia (FA) predisposes to hematopoietic failure, birth defects, leukemia, and squamous cell carcinoma of the head and neck (HNSCC) and cervix. The FA/BRCA pathway includes 8 members of a core complex and 5 downstream gene products closely linked with BRCA1 or BRCA2. Precancerous lesions are believed to trigger the DNA damage response (DDR), and we focused on the DDR in FA and its putative role as a checkpoint barrier to cancer. In primary fibroblasts with mutations in the core complex FANCA protein, we discovered that basal expression and phosphorylation of ATM (ataxia telangiectasia mutated) and p53 induced by irradiation (IR) or mitomycin C (MMC) were upregulated. This heightened response appeared to be due to increased basal levels of ATM in cultured FANCA-mutant cells, highlighting the new observation that ATM can be regulated at the transcriptional level in addition to its well-established activation by autophosphorylation. Functional analysis of this response using gamma-H2AX foci as markers of DNA double-stranded breaks (DSBs) demonstrated abnormal persistence of only MMC- and not IR-induced foci. Thus, we describe a processing defect that leads to general DDR upregulation but specific persistence of DNA crosslinker-induced damage response foci. Underscoring the significance of these findings, we found resistance to DNA crosslinker-induced cell cycle arrest and apoptosis in a TP53-mutant, patient-derived HNSCC cell line, whereas a lymphoblastoid cell line derived from this same individual was not mutated at TP53 and retained DNA crosslinker sensitivity. Our results suggest that cancer in FA may arise from selection for cells that escape from a chronically activated DDR checkpoint.

  4. Toxic effect of silica nanoparticles on endothelial cells through DNA damage response via Chk1-dependent G2/M checkpoint.

    Directory of Open Access Journals (Sweden)

    Junchao Duan

    Full Text Available Silica nanoparticles have become promising carriers for drug delivery or gene therapy. Endothelial cells could be directly exposed to silica nanoparticles by intravenous administration. However, the underlying toxic effect mechanisms of silica nanoparticles on endothelial cells are still poorly understood. In order to clarify the cytotoxicity of endothelial cells induced by silica nanoparticles and its mechanisms, cellular morphology, cell viability and lactate dehydrogenase (LDH release were observed in human umbilical vein endothelial cells (HUVECs as assessing cytotoxicity, resulted in a dose- and time- dependent manner. Silica nanoparticles-induced reactive oxygen species (ROS generation caused oxidative damage followed by the production of malondialdehyde (MDA as well as the inhibition of superoxide dismutase (SOD and glutathione peroxidase (GSH-Px. Both necrosis and apoptosis were increased significantly after 24 h exposure. The mitochondrial membrane potential (MMP decreased obviously in a dose-dependent manner. The degree of DNA damage including the percentage of tail DNA, tail length and Olive tail moment (OTM were markedly aggravated. Silica nanoparticles also induced G2/M arrest through the upregulation of Chk1 and the downregulation of Cdc25C, cyclin B1/Cdc2. In summary, our data indicated that the toxic effect mechanisms of silica nanoparticles on endothelial cells was through DNA damage response (DDR via Chk1-dependent G2/M checkpoint signaling pathway, suggesting that exposure to silica nanoparticles could be a potential hazards for the development of cardiovascular diseases.

  5. Histone dosage regulates DNA damage sensitivity in a checkpoint-independent manner by the homologous recombination pathway

    Science.gov (United States)

    Liang, Dun; Burkhart, Sarah Lyn; Singh, Rakesh Kumar; Kabbaj, Marie-Helene Miquel; Gunjan, Akash

    2012-01-01

    In eukaryotes, multiple genes encode histone proteins that package genomic deoxyribonucleic acid (DNA) and regulate its accessibility. Because of their positive charge, ‘free’ (non-chromatin associated) histones can bind non-specifically to the negatively charged DNA and affect its metabolism, including DNA repair. We have investigated the effect of altering histone dosage on DNA repair in budding yeast. An increase in histone gene dosage resulted in enhanced DNA damage sensitivity, whereas deletion of a H3–H4 gene pair resulted in reduced levels of free H3 and H4 concomitant with resistance to DNA damaging agents, even in mutants defective in the DNA damage checkpoint. Studies involving the repair of a HO endonuclease-mediated DNA double-strand break (DSB) at the MAT locus show enhanced repair efficiency by the homologous recombination (HR) pathway on a reduction in histone dosage. Cells with reduced histone dosage experience greater histone loss around a DSB, whereas the recruitment of HR factors is concomitantly enhanced. Further, free histones compete with the HR machinery for binding to DNA and associate with certain HR factors, potentially interfering with HR-mediated repair. Our findings may have important implications for DNA repair, genomic stability, carcinogenesis and aging in human cells that have dozens of histone genes. PMID:22850743

  6. Loss of yeast peroxiredoxin Tsa1p induces genome instability through activation of the DNA damage checkpoint and elevation of dNTP levels.

    Directory of Open Access Journals (Sweden)

    Hei-Man Vincent Tang

    2009-10-01

    Full Text Available Peroxiredoxins are a family of antioxidant enzymes critically involved in cellular defense and signaling. Particularly, yeast peroxiredoxin Tsa1p is thought to play a role in the maintenance of genome integrity, but the underlying mechanism is not understood. In this study, we took a genetic approach to investigate the cause of genome instability in tsa1Delta cells. Strong genetic interactions of TSA1 with DNA damage checkpoint components DUN1, SML1, and CRT1 were found when mutant cells were analyzed for either sensitivity to DNA damage or rate of spontaneous base substitutions. An elevation in intracellular dNTP production was observed in tsa1Delta cells. This was associated with constitutive activation of the DNA damage checkpoint as indicated by phosphorylation of Rad9/Rad53p, reduced steady-state amount of Sml1p, and induction of RNR and HUG1 genes. In addition, defects in the DNA damage checkpoint did not modulate intracellular level of reactive oxygen species, but suppressed the mutator phenotype of tsa1Delta cells. On the contrary, overexpression of RNR1 exacerbated this phenotype by increasing dNTP levels. Taken together, our findings uncover a new role of TSA1 in preventing the overproduction of dNTPs, which is a root cause of genome instability.

  7. C/EBPα Expression is Partially Regulated by C/EBPβ in Response to DNA Damage and C/EBPα Deficient Fibroblasts Display an Impaired G1 Checkpoint

    Science.gov (United States)

    Ranjan, Rakesh; Thompson, Elizabeth A.; Yoon, Kyungsil; Smart, Robert C.

    2009-01-01

    We observed that C/EBPα is highly inducible in primary fibroblasts by DNA damaging agents that induce strand breaks, alkylate and crosslink DNA as well as those that produce bulky DNA lesions. Fibroblasts deficient in C/EBPα (C/EBPα-/-) display an impaired G1 checkpoint as evidenced by inappropriate entry into S-phase in response to DNA damage and these cells also display an enhanced G1 to S transition in response to mitogens. The induction of C/EBPα by DNA damage in fibroblasts does not require p53. EMSA analysis of nuclear extracts prepared from UVB- and MNNG-treated fibroblasts revealed increased binding of C/EBPβ to a C/EBP consensus sequence and ChIP analysis revealed increased C/EBPβ binding to the C/EBPα promoter. To determine whether C/EBPβ has a role in the regulation of C/EBPα we treated C/EBPβ-/- fibroblasts with UVB or MNNG. We observed C/EBPα induction was impaired in both UVB- and MNNG- treated C/EBPβ-/- fibroblasts. Our study reveals a novel role for C/EBPβ in the regulation of C/EBPα in response to DNA damage and provides definitive genetic evidence that C/EBPα has a critical role in the DNA damage G1 checkpoint. PMID:19581927

  8. The Aurora-B-dependent NoCut checkpoint prevents damage of anaphase bridges after DNA replication stress.

    Science.gov (United States)

    Amaral, Nuno; Vendrell, Alexandre; Funaya, Charlotta; Idrissi, Fatima-Zahra; Maier, Michael; Kumar, Arun; Neurohr, Gabriel; Colomina, Neus; Torres-Rosell, Jordi; Geli, María-Isabel; Mendoza, Manuel

    2016-05-01

    Anaphase chromatin bridges can lead to chromosome breakage if not properly resolved before completion of cytokinesis. The NoCut checkpoint, which depends on Aurora B at the spindle midzone, delays abscission in response to chromosome segregation defects in yeast and animal cells. How chromatin bridges are detected, and whether abscission inhibition prevents their damage, remain key unresolved questions. We find that bridges induced by DNA replication stress and by condensation or decatenation defects, but not dicentric chromosomes, delay abscission in a NoCut-dependent manner. Decatenation and condensation defects lead to spindle stabilization during cytokinesis, allowing bridge detection by Aurora B. NoCut does not prevent DNA damage following condensin or topoisomerase II inactivation; however, it protects anaphase bridges and promotes cellular viability after replication stress. Therefore, the molecular origin of chromatin bridges is critical for activation of NoCut, which plays a key role in the maintenance of genome stability after replicative stress.

  9. The G2/M DNA damage checkpoint inhibits mitosis through Tyr15 phosphorylation of p34cdc2 in Aspergillus nidulans.

    Science.gov (United States)

    Ye, X S; Fincher, R R; Tang, A; Osmani, S A

    1997-01-02

    It is possible to cause G2 arrest in Aspergillus nidulans by inactivating either p34cdc2 or NIMA. We therefore investigated the negative control of these two mitosis-promoting kinases after DNA damage. DNA damage caused rapid Tyr15 phosphorylation of p34cdc2 and transient cell cycle arrest but had little effect on the activity of NIMA. Dividing cells deficient in Tyr15 phosphorylation of p34cdc2 were sensitive to both MMS and UV irradiation and entered lethal premature mitosis with damaged DNA. However, non-dividing quiescent conidiospores of the Tyr15 mutant strain were not sensitive to DNA damage. The UV and MMS sensitivity of cells unable to tyrosine phosphorylate p34cdc2 is therefore caused by defects in DNA damage checkpoint regulation over mitosis. Both the nimA5 and nimT23 temperature-sensitive mutations cause an arrest in G2 at 42 degrees C. Addition of MMS to nimT23 G2-arrested cells caused a marked delay in their entry into mitosis upon downshift to 32 degrees C and this delay was correlated with a long delay in the dephosphorylation and activation of p34cdc2. Addition of MMS to nimA5 G2-arrested cells caused inactivation of the H1 kinase activity of p34cdc2 due to an increase in its Tyr15 phosphorylation level and delayed entry into mitosis upon return to 32 degrees C. However, if Tyr15 phosphorylation of p34cdc2 was prevented then its H1 kinase activity was not inactivated upon MMS addition to nimA5 G2-arrested cells and they rapidly progressed into a lethal mitosis upon release to 32 degrees C. Thus, Tyr15 phosphorylation of p34cdc2 in G2 arrests initiation of mitosis after DNA damage in A. nidulans.

  10. The co-repressor SMRT delays DNA damage-induced caspase activation by repressing pro-apoptotic genes and modulating the dynamics of checkpoint kinase 2 activation.

    Directory of Open Access Journals (Sweden)

    Claudio Scafoglio

    Full Text Available Checkpoint kinase 2 (Chk2 is a major regulator of DNA damage response and can induce alternative cellular responses: cell cycle arrest and DNA repair or programmed cell death. Here, we report the identification of a new role of Chk2 in transcriptional regulation that also contributes to modulating the balance between survival and apoptosis following DNA damage. We found that Chk2 interacts with members of the NCoR/SMRT transcriptional co-regulator complexes and serves as a functional component of the repressor complex, being required for recruitment of SMRT on the promoter of pro-apoptotic genes upon DNA damage. Thus, the co-repressor SMRT exerts a critical protective action against genotoxic stress-induced caspase activation, repressing a functionally important cohort of pro-apoptotic genes. Amongst them, SMRT is responsible for basal repression of Wip1, a phosphatase that de-phosphorylates and inactivates Chk2, thus affecting a feedback loop responsible for licensing the correct timing of Chk2 activation and the proper execution of the DNA repair process.

  11. Stimulation of functional vision in children with perinatal brain damage.

    Science.gov (United States)

    Alimović, Sonja; Mejaski-Bosnjak, Vlatka

    2011-01-01

    Cerebral visual impairment (CVI) is one of the most common causes of bilateral visual loss, which frequently occurs due to perinatal brain injury. Vision in early life has great impact on acquisition of basic comprehensions which are fundamental for further development. Therefore, early detection of visual problems and early intervention is necessary. The aim of the present study is to determine specific visual functioning of children with perinatal brain damage and the influence of visual stimulation on development of functional vision at early age of life. We initially assessed 30 children with perinatal brain damage up to 3 years of age, who were reffered to our pediatric low vision cabinet in "Little house" from child neurologists, ophthalmologists Type and degree of visual impairment was determined according to functional vision assessment of each child. On the bases of those assessments different kind of visual stimulations were carried out with children who have been identified to have a certain visual impairment. Through visual stimulation program some of the children were stimulated with light stimulus, some with different materials under the ultraviolet (UV) light, and some with bright color and high contrast materials. Children were also involved in program of early stimulation of overall sensory motor development. Goals and methods of therapy were determined individually, based on observation of child's possibilities and need. After one year of program, reassessment was done. Results for visual functions and functional vision were compared to evaluate the improvement of the vision development. These results have shown that there was significant improvement in functional vision, especially in visual attention and visual communication.

  12. C/EBPα regulates CRL4Cdt2-mediated degradation of p21 in response to UVB-induced DNA damage to control the G1/S checkpoint

    Science.gov (United States)

    Hall, Jonathan R; Bereman, Michael S; Nepomuceno, Angelito I; Thompson, Elizabeth A; Muddiman, David C; Smart, Robert C

    2014-01-01

    The bZIP transcription factor, C/EBPα is highly inducible by UVB and other DNA damaging agents in keratinocytes. C/EBPα-deficient keratinocytes fail to undergo cell cycle arrest in G1 in response to UVB-induced DNA damage and mice lacking epidermal C/EBPα are highly susceptible to UVB-induced skin cancer. The mechanism through which C/EBPα regulates the cell cycle checkpoint in response to DNA damage is unknown. Here we report untreated C/EBPα-deficient keratinocytes have normal levels of the cyclin-dependent kinase inhibitor, p21, however, UVB-treated C/EBPα-deficient keratinocytes fail to up-regulate nuclear p21 protein levels despite normal up-regulation of Cdkn1a mRNA levels. UVB-treated C/EBPα-deficient keratinocytes displayed a 4-fold decrease in nuclear p21 protein half-life due to the increased proteasomal degradation of p21 via the E3 ubiquitin ligase CRL4Cdt2. Cdt2 is the substrate recognition subunit of CRL4Cdt2 and Cdt2 mRNA and protein levels were up-regulated in UVB-treated C/EBPα-deficient keratinocytes. Knockdown of Cdt2 restored p21 protein levels in UVB-treated C/EBPα-deficient keratinocytes. Lastly, the failure to accumulate p21 in response to UVB in C/EBPα-deficient keratinocytes resulted in decreased p21 interactions with critical cell cycle regulatory proteins, increased CDK2 activity, and inappropriate entry into S-phase. These findings reveal C/EBPα regulates G1/S cell cycle arrest in response to DNA damage via the control of CRL4Cdt2 mediated degradation of p21. PMID:25483090

  13. Rationale for combination of therapeutic antibodies targeting tumor cells and immune checkpoint receptors: Harnessing innate and adaptive immunity through IgG1 isotype immune effector stimulation.

    Science.gov (United States)

    Ferris, Robert L; Lenz, Heinz-Josef; Trotta, Anna Maria; García-Foncillas, Jesús; Schulten, Jeltje; Audhuy, François; Merlano, Marco; Milano, Gerard

    2018-02-01

    Immunoglobulin (Ig) G1 antibodies stimulate antibody-dependent cell-mediated cytotoxicity (ADCC). Cetuximab, an IgG1 isotype monoclonal antibody, is a standard-of-care treatment for locally advanced and recurrent and/or metastatic squamous cell carcinoma of the head and neck (SCCHN) and metastatic colorectal cancer (CRC). Here we review evidence regarding the clinical relevance of cetuximab-mediated ADCC and other immune functions and provide a biological rationale concerning why this property positions cetuximab as an ideal partner for immune checkpoint inhibitors (ICIs) and other emerging immunotherapies. We performed a nonsystematic review of available preclinical and clinical data involving cetuximab-mediated immune activity and combination approaches of cetuximab with other immunotherapies, including ICIs, in SCCHN and CRC. Indeed, cetuximab mediates ADCC activity in the intratumoral space and primes adaptive and innate cellular immunity. However, counterregulatory mechanisms may lead to immunosuppressive feedback loops. Accordingly, there is a strong rationale for combining ICIs with cetuximab for the treatment of advanced tumors, as targeting CTLA-4, PD-1, and PD-L1 can ostensibly overcome these immunosuppressive counter-mechanisms in the tumor microenvironment. Moreover, combining ICIs (or other immunotherapies) with cetuximab is a promising strategy for boosting immune response and enhancing response rates and durability of response. Cetuximab immune activity-including, but not limited to, ADCC-provides a strong rationale for its combination with ICIs or other immunotherapies to synergistically and fully mobilize the adaptive and innate immunity against tumor cells. Ongoing prospective studies will evaluate the clinical effect of these combination regimens and their immune effect in CRC and SCCHN and in other indications. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. A plant cyclin B2 is degraded early in mitosis and its ectopic expression shortens G2-phase and alleviates the DNA-damage checkpoint

    Czech Academy of Sciences Publication Activity Database

    Weingartner, M.; Pelayo, H. R.; Binarová, Pavla; Zwerger, K.; Melikant, B.; Torre, C.; Heberle-Bors, E.; Bogre, L.

    2003-01-01

    Roč. 116, č. 3 (2003), s. 487-498 ISSN 0021-9533 R&D Projects: GA MŠk LN00A081 Grant - others:GA Ministerio de Ciencia y Tecnologia(ES) BMC2001-2195; Biotechnology and Biological Science Research Council(AU) 111/P133340 Institutional research plan: CEZ:AV0Z5020903 Keywords : cyclin B * mitosis * checkpoint Subject RIV: EE - Microbiology, Virology Impact factor: 7.250, year: 2003

  15. "Isogaba Maware": quality control of genome DNA by checkpoints.

    Science.gov (United States)

    Kitazono, A; Matsumoto, T

    1998-05-01

    Checkpoints maintain the interdependency of cell cycle events by permitting the onset of an event only after the completion of the preceding event. The DNA replication checkpoint induces a cell cycle arrest until the completion of the DNA replication. Similarly, the DNA damage checkpoint arrests cell cycle progression if DNA repair is incomplete. A number of genes that play a role in the two checkpoints have been identified through genetic studies in yeasts, and their homologues have been found in fly, mouse, and human. They form signaling cascades activated by a DNA replication block or DNA damage and subsequently generate the negative constraints on cell cycle regulators. The failure of these signaling cascades results in producing offspring that carry mutations or that lack a portion of the genome. In humans, defects in the checkpoints are often associated with cancer-prone diseases. Focusing mainly on the studies in budding and fission yeasts, we summarize the recent progress.

  16. Targeting the Checkpoint to Kill Cancer Cells

    Czech Academy of Sciences Publication Activity Database

    Benada, Jan; Macůrek, Libor

    2015-01-01

    Roč. 6, č. 3 (2015), s. 1912-1937 ISSN 2218-273X R&D Projects: GA ČR(CZ) GA14-34264S Institutional support: RVO:68378050 Keywords : checkpoint * DNA damage response * cancer Subject RIV: EB - Genetics ; Molecular Biology

  17. Stimulation of Functional Vision in Children with Perinatal Brain Damage

    OpenAIRE

    Alimović, Sonja; Mejaški-Bošnjak, Vlatka

    2011-01-01

    Cerebral visual impairment (CVI) is one of the most common causes of bilateral visual loss, which frequently occurs due to perinatal brain injury. Vision in early life has great impact on acquisition of basic comprehensions which are fundamental for further development. Therefore, early detection of visual problems and early intervention is necessary. The aim of the present study is to determine specific visual functioning of children with perinatal brain damage and the influence of visual st...

  18. Recreational stimulants, herbal, and spice cannabis: The core psychobiological processes that underlie their damaging effects.

    Science.gov (United States)

    Parrott, Andrew C; Hayley, Amie C; Downey, Luke A

    2017-05-01

    Recreational drugs are taken for their positive mood effects, yet their regular usage damages well-being. The psychobiological mechanisms underlying these damaging effects will be debated. The empirical literature on recreational cannabinoids and stimulant drugs is reviewed. A theoretical explanation for how they cause similar types of damage is outlined. All psychoactive drugs cause moods and psychological states to fluctuate. The acute mood gains underlie their recreational usage, while the mood deficits on withdrawal explain their addictiveness. Cyclical mood changes are found with every central nervous system stimulant and also occur with cannabis. These mood state changes provide a surface index for more profound psychobiological fluctuations. Homeostatic balance is altered, with repetitive disturbances of the hypothalamic-pituitary-adrenal axis, and disrupted cortisol-neurohormonal secretions. Hence, these drugs cause increased stress, disturbed sleep, neurocognitive impairments, altered brain activity, and psychiatric vulnerability. Equivalent deficits occur with novel psychoactive stimulants such as mephedrone and artificial "spice" cannabinoids. These psychobiological fluctuations underlie drug dependency and make cessation difficult. Psychobiological stability and homeostatic balance are optimally restored by quitting psychoactive drugs. Recreational stimulants such as cocaine or MDMA (3.4-methylenedioxymethamphetamine) and sedative drugs such as cannabis damage human homeostasis and well-being through similar core psychobiological mechanisms. Copyright © 2017 John Wiley & Sons, Ltd.

  19. Combination approaches with immune checkpoint blockade in cancer therapy

    Directory of Open Access Journals (Sweden)

    Maarten Swart

    2016-11-01

    Full Text Available In healthy individuals, immune checkpoint molecules prevent autoimmune responses and limit immune cell-mediated tissue damage. Tumors frequently exploit these molecules to evade eradication by the immune system. Over the past years, immune checkpoint blockade of cytotoxic T lymphocyte antigen-4 (CTLA-4 and programmed death-1 (PD-1 emerged as promising strategies to activate anti-tumor cytotoxic T cell responses. Although complete regression and long-term survival is achieved in some patients, not all patients respond. This review describes promising, novel combination approaches involving immune checkpoint blockade, aimed at increasing response-rates to the single treatments.

  20. The G2/M DNA damage checkpoint inhibits mitosis through Tyr15 phosphorylation of p34cdc2 in Aspergillus nidulans.

    OpenAIRE

    Ye, X S; Fincher, R R; Tang, A; Osmani, S A

    1997-01-01

    It is possible to cause G2 arrest in Aspergillus nidulans by inactivating either p34cdc2 or NIMA. We therefore investigated the negative control of these two mitosis-promoting kinases after DNA damage. DNA damage caused rapid Tyr15 phosphorylation of p34cdc2 and transient cell cycle arrest but had little effect on the activity of NIMA. Dividing cells deficient in Tyr15 phosphorylation of p34cdc2 were sensitive to both MMS and UV irradiation and entered lethal premature mitosis with damaged DN...

  1. F-box protein FBXO31 is a dedicated checkpoint protein to facilitate cell cycle arrest through activation of regulators in radiation induced DNA damage

    International Nuclear Information System (INIS)

    Santra, Manas Kumar

    2017-01-01

    In response to radiation-induced DNA damage, eukaryotic cells initiate a complex signalling pathway, termed the DNA damage response (DDR), which coordinates cell cycle arrest with DNA repair. Previous study showed that induction of G1 arrest in response to radiation induced DNA damage is minimally a two-step process: a fast p53-independent initiation of G1 arrest mediated by cyclin D1 proteolysis and a slower maintenance of arrest resulting from increased p53 stability. We elucidated the molecular mechanism of slow and fast response of radiation induced DDR. We showed that FBXO31, a member of F-box family proteins, plays important role in DDR induced by ionizing radiation. We show that FBXO31 is responsible for promoting MDM2 degradation following radiation. FBXO31 interacts with and directs the degradation of MDM2 in ATM dependent phosphorylation of MDM2. FBXO31-mediated loss of MDM2 leads to elevated levels of p53, resulting in growth arrest. In cells depleted of FBXO31, MDM2 is not degraded and p53 levels do not increase following genotoxic stress. Thus, FBXO31 is essential for the classic robust increase in p53 levels following DNA damage

  2. Checkpoint independence of most DNA replication origins in fission yeast.

    Science.gov (United States)

    Mickle, Katie L; Ramanathan, Sunita; Rosebrock, Adam; Oliva, Anna; Chaudari, Amna; Yompakdee, Chulee; Scott, Donna; Leatherwood, Janet; Huberman, Joel A

    2007-12-19

    -mutant cells. The fact that approximately 97% of fission yeast replication origins - both early and late - are not significantly affected by replication checkpoint mutations in HU-treated cells suggests that (i) most late-firing origins are restrained from firing in HU-treated cells by at least one checkpoint-independent mechanism, and (ii) checkpoint-dependent slowing of S phase in fission yeast when DNA is damaged may be accomplished primarily by the slowing of replication forks.

  3. Checkpoint independence of most DNA replication origins in fission yeast

    Science.gov (United States)

    Mickle, Katie L; Ramanathan, Sunita; Rosebrock, Adam; Oliva, Anna; Chaudari, Amna; Yompakdee, Chulee; Scott, Donna; Leatherwood, Janet; Huberman, Joel A

    2007-01-01

    checkpoint-mutant cells. Conclusion The fact that ~97% of fission yeast replication origins – both early and late – are not significantly affected by replication checkpoint mutations in HU-treated cells suggests that (i) most late-firing origins are restrained from firing in HU-treated cells by at least one checkpoint-independent mechanism, and (ii) checkpoint-dependent slowing of S phase in fission yeast when DNA is damaged may be accomplished primarily by the slowing of replication forks. PMID:18093330

  4. Checkpoint independence of most DNA replication origins in fission yeast

    Directory of Open Access Journals (Sweden)

    Scott Donna

    2007-12-01

    -type cells as in checkpoint-mutant cells. Conclusion The fact that ~97% of fission yeast replication origins – both early and late – are not significantly affected by replication checkpoint mutations in HU-treated cells suggests that (i most late-firing origins are restrained from firing in HU-treated cells by at least one checkpoint-independent mechanism, and (ii checkpoint-dependent slowing of S phase in fission yeast when DNA is damaged may be accomplished primarily by the slowing of replication forks.

  5. The spindle assembly checkpoint: More than just keeping track of the spindle.

    OpenAIRE

    Lawrence, KS; Engebrecht, J

    2015-01-01

    Genome stability is essential for cell proliferation and survival. Consequently, genome integrity is monitored by two major checkpoints, the DNA damage response (DDR) and the spindle assembly checkpoint (SAC). The DDR monitors DNA lesions in G1, S, and G2 stages of the cell cycle and the SAC ensures proper chromosome segregation in M phase. There have been extensive studies characterizing the roles of these checkpoints in response to the processes for which they are named; however, emerging e...

  6. Nonlinear ultrasonic stimulated thermography for damage assessment in isotropic fatigued structures

    Science.gov (United States)

    Fierro, Gian Piero Malfense; Calla', Danielle; Ginzburg, Dmitri; Ciampa, Francesco; Meo, Michele

    2017-09-01

    Traditional non-destructive evaluation (NDE) and structural health monitoring (SHM) systems are used to analyse that a structure is free of any harmful damage. However, these techniques still lack sensitivity to detect the presence of material micro-flaws in the form of fatigue damage and often require time-consuming procedures and expensive equipment. This research work presents a novel "nonlinear ultrasonic stimulated thermography" (NUST) method able to overcome some of the limitations of traditional linear ultrasonic/thermography NDE-SHM systems and to provide a reliable, rapid and cost effective estimation of fatigue damage in isotropic materials. Such a hybrid imaging approach combines the high sensitivity of nonlinear acoustic/ultrasonic techniques to detect micro-damage, with local defect frequency selection and infrared imaging. When exciting structures with an optimised frequency, nonlinear elastic waves are observed and higher frictional work at the fatigue damaged area is generated due to clapping and rubbing of the crack faces. This results in heat at cracked location that can be measured using an infrared camera. A Laser Vibrometer (LV) was used to evaluate the extent that individual frequency components contribute to the heating of the damage region by quantifying the out-of-plane velocity associated with the fundamental and second order harmonic responses. It was experimentally demonstrated the relationship between a nonlinear ultrasound parameter (βratio) of the material nonlinear response to the actual temperature rises near the crack. These results demonstrated that heat generation at damaged regions could be amplified by exciting at frequencies that provide nonlinear responses, thus improving the imaging of material damage and the reliability of NUST in a quick and reproducible manner.

  7. Differences in the stimulation of repair replication by 3-aminobenzamide in lymphoblastoid cells damaged by methylmethanesulfonate or ultraviolet light

    Energy Technology Data Exchange (ETDEWEB)

    Cleaver, J.E.; Morgan, W.F.

    1987-09-01

    Human lymphoblastoid cells damaged by u.v. light accumulated DNA breaks in the presence of cytosine arabinoside and hydroxyurea at a frequency similar to that of cells damaged by methylmethanesulfonate. 3-Aminobenzamide (1 mM) reduced the net strand-break frequency detected after either kind of damage. Repair replication, however, was stimulated only in methylmethanesulfonate-damaged cells. This stimulation is therefore not related directly to the DNA strand-break frequencies and concomitant poly(ADP-ribose) synthesis, but depends on some other cellular response specific to alkylating agents.

  8. A genetic screen identifies BRCA2 and PALB2 as key regulators of G2 checkpoint maintenance

    DEFF Research Database (Denmark)

    Menzel, Tobias; Nähse-Kumpf, Viola; Kousholt, Arne Nedergaard

    2011-01-01

    To identify key connections between DNA-damage repair and checkpoint pathways, we performed RNA interference screens for regulators of the ionizing radiation-induced G2 checkpoint, and we identified the breast cancer gene BRCA2. The checkpoint was also abrogated following depletion of PALB2......, an interaction partner of BRCA2. BRCA2 and PALB2 depletion led to premature checkpoint abrogation and earlier activation of the AURORA A-PLK1 checkpoint-recovery pathway. These results indicate that the breast cancer tumour suppressors and homologous recombination repair proteins BRCA2 and PALB2 are main...

  9. Using LEDs to stimulate the recovery of radiation damage to plastic scintillators

    Energy Technology Data Exchange (ETDEWEB)

    Wetzel, J., E-mail: james-wetzel@uiowa.edu [The University of Iowa, Iowa City, IA (United States); Tiras, E. [The University of Iowa, Iowa City, IA (United States); Bilki, B. [The University of Iowa, Iowa City, IA (United States); Beykent University, Istanbul (Turkey); Onel, Y. [The University of Iowa, Iowa City, IA (United States); Winn, D. [Fairfield University, Fairfield, CT (United States)

    2017-03-15

    In this study, we consider using LEDs to stimulate the recovery of scintillators damaged from radiation in high radiation environments. We irradiated scintillating tiles of polyethylene naphthalate (PEN), Eljen brand EJ-260 (EJN), an overdoped EJ-260 (EJ2P), and a lab-produced elastomer scintillator (ES) composed of p-terphenyl (ptp) in epoxy. Two different high-dose irradiations took place, with PEN dosed to 100 kGy, and the others to 78 kGy. We found that the ‘blue’ scintillators (PEN and ES) recovered faster and maximally higher with LEDs than without. Conversely exposing the ‘green’ scintillators (EJ-260) to LED light had a nearly negligible effect on the recovery. We hypothesize that the ‘green’ scintillators require wavelengths that match their absorption and emission spectra for LED stimulated recovery.

  10. Use of lectin-induced lymphocyte stimulation as a biodosimeter of radiation damage

    International Nuclear Information System (INIS)

    Shifrine, M.; Taylor, N.J.; DeRock, E.W.; Wiger, N.; Wilson, F.D.

    1979-01-01

    The purpose of this study was to determine the feasibility of utilizing an in vitro test to determine whether an individual has suffered radiation damage. It was not our purpose to develop a test capable of detecting low doses but rather to determine possible damage due to a radiation accident at medium to high doses. In a pilot study using the whole blood lymphocyte stimulation test (WB/LST), we pretested six dogs weekly for three weeks. One was sham irradiated and is referred to as the control and one each received an acute whole body dose of 10, 25, 50, 75, or 100 R. The dogs were tested one day post-irradiation and then weekly for 56 days. Our data indicate that, using Con-A and PHA in the WB/LST, a biodosimeter can be developed to determine recovery of lymphocyte function after suspected radiation exposure to confirm accidental radiation and estimate the magnitude of the dose

  11. Phosphorylation of Minichromosome Maintenance 3 (MCM3) by Checkpoint Kinase 1 (Chk1) Negatively Regulates DNA Replication and Checkpoint Activation.

    Science.gov (United States)

    Han, Xiangzi; Mayca Pozo, Franklin; Wisotsky, Jacob N; Wang, Benlian; Jacobberger, James W; Zhang, Youwei

    2015-05-08

    Mechanisms controlling DNA replication and replication checkpoint are critical for the maintenance of genome stability and the prevention or treatment of human cancers. Checkpoint kinase 1 (Chk1) is a key effector protein kinase that regulates the DNA damage response and replication checkpoint. The heterohexameric minichromosome maintenance (MCM) complex is the core component of mammalian DNA helicase and has been implicated in replication checkpoint activation. Here we report that Chk1 phosphorylates the MCM3 subunit of the MCM complex at Ser-205 under normal growth conditions. Mutating the Ser-205 of MCM3 to Ala increased the length of DNA replication track and shortened the S phase duration, indicating that Ser-205 phosphorylation negatively controls normal DNA replication. Upon replicative stress treatment, the inhibitory phosphorylation of MCM3 at Ser-205 was reduced, and this reduction was accompanied with the generation of single strand DNA, the key platform for ataxia telangiectasia mutated and Rad3-related (ATR) activation. As a result, the replication checkpoint is activated. Together, these data provide significant insights into the regulation of both normal DNA replication and replication checkpoint activation through the novel phosphorylation of MCM3 by Chk1. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Cell cycle checkpoints: reversible when possible, irreversible when needed

    NARCIS (Netherlands)

    Krenning, L.

    2015-01-01

    Cell cycle checkpoints are reversible in nature, and can prevent progression into the next cell cycle phase if needed. In the case of DNA damage, cells can prevent progression from G1 into S phase, and from G2 into mitosis in the presence of DNA double strand breaks. Following DNA repair, these

  13. Checkpoint responses to replication stalling: inducing tolerance and preventing mutagenesis

    Energy Technology Data Exchange (ETDEWEB)

    Kai, Mihoko; Wang, Teresa S.-F

    2003-11-27

    Replication mutants often exhibit a mutator phenotype characterized by point mutations, single base frameshifts, and the deletion or duplication of sequences flanked by homologous repeats. Mutation in genes encoding checkpoint proteins can significantly affect the mutator phenotype. Here, we use fission yeast (Schizosaccharomyces pombe) as a model system to discuss the checkpoint responses to replication perturbations induced by replication mutants. Checkpoint activation induced by a DNA polymerase mutant, aside from delay of mitotic entry, up-regulates the translesion polymerase DinB (Pol{kappa}). Checkpoint Rad9-Rad1-Hus1 (9-1-1) complex, which is loaded onto chromatin by the Rad17-Rfc2-5 checkpoint complex in response to replication perturbation, recruits DinB onto chromatin to generate the point mutations and single nucleotide frameshifts in the replication mutator. This chain of events reveals a novel checkpoint-induced tolerance mechanism that allows cells to cope with replication perturbation, presumably to make possible restarting stalled replication forks. Fission yeast Cds1 kinase plays an essential role in maintaining DNA replication fork stability in the face of DNA damage and replication fork stalling. Cds1 kinase is known to regulate three proteins that are implicated in maintaining replication fork stability: Mus81-Eme1, a hetero-dimeric structure-specific endonuclease complex; Rqh1, a RecQ-family helicase involved in suppressing inappropriate recombination during replication; and Rad60, a protein required for recombinational repair during replication. These Cds1-regulated proteins are thought to cooperatively prevent mutagenesis and maintain replication fork stability in cells under replication stress. These checkpoint-regulated processes allow cells to survive replication perturbation by preventing stalled replication forks from degenerating into deleterious DNA structures resulting in genomic instability and cancer development.

  14. Checkpoint responses to replication stalling: inducing tolerance and preventing mutagenesis

    International Nuclear Information System (INIS)

    Kai, Mihoko; Wang, Teresa S.-F.

    2003-01-01

    Replication mutants often exhibit a mutator phenotype characterized by point mutations, single base frameshifts, and the deletion or duplication of sequences flanked by homologous repeats. Mutation in genes encoding checkpoint proteins can significantly affect the mutator phenotype. Here, we use fission yeast (Schizosaccharomyces pombe) as a model system to discuss the checkpoint responses to replication perturbations induced by replication mutants. Checkpoint activation induced by a DNA polymerase mutant, aside from delay of mitotic entry, up-regulates the translesion polymerase DinB (Polκ). Checkpoint Rad9-Rad1-Hus1 (9-1-1) complex, which is loaded onto chromatin by the Rad17-Rfc2-5 checkpoint complex in response to replication perturbation, recruits DinB onto chromatin to generate the point mutations and single nucleotide frameshifts in the replication mutator. This chain of events reveals a novel checkpoint-induced tolerance mechanism that allows cells to cope with replication perturbation, presumably to make possible restarting stalled replication forks. Fission yeast Cds1 kinase plays an essential role in maintaining DNA replication fork stability in the face of DNA damage and replication fork stalling. Cds1 kinase is known to regulate three proteins that are implicated in maintaining replication fork stability: Mus81-Eme1, a hetero-dimeric structure-specific endonuclease complex; Rqh1, a RecQ-family helicase involved in suppressing inappropriate recombination during replication; and Rad60, a protein required for recombinational repair during replication. These Cds1-regulated proteins are thought to cooperatively prevent mutagenesis and maintain replication fork stability in cells under replication stress. These checkpoint-regulated processes allow cells to survive replication perturbation by preventing stalled replication forks from degenerating into deleterious DNA structures resulting in genomic instability and cancer development

  15. ATM and checkpoint responses to DNA double strand breaks

    International Nuclear Information System (INIS)

    Khanna, K.K.

    2003-01-01

    DNA damage checkpoints can be classified into G1/S, intra-S and G2/M checkpoints, so named according to the cell cycle transitions that they regulate. DNA damage incurred during the G1 or G2 phase of the cell cycle leads to growth arrest at the G1/S and G2/M phase boundaries, respectively, whereas genotoxic stress during S phase results in the transient suppression of DNA synthesis. In mammals, ATM (ataxia-telangiectasia mutated) is a protein kinase that controls all checkpoint responses to DNA damage. ATM is a versatile kinase which uses various means to regulate a given checkpoint pathway. It has been shown to act upon several proteins within the same pathway, many times controlling several different modifications of the same protein or using several different targets to arrive at the same end point. Some of the ATM targets act as adaptors by recruiting additional substrates for ATM. ATM controls two types of responses in G1. The p53-dependent responses inhibit Cyclin/Cdk activity by transcriptional induction of p21, whereas p53-independent responses inhibit CDKs through degradation of Cdc25A to maintain CdK2 inhibitory phosphorylation. In regulating p53, ATM directly phosphorylates p53 on Ser15, which likely causes p53 transcriptional activation, concurrently activating other kinases that phosphorylate p53 at other sites such as Ser20, which reduces the ability of MDM2 to bind p53, thus promoting its stability. ATM further ensures p53 stability by phosphorylating MDM2. At least six ATM targets, namely CHK2, CHK1, NBS1, BRCA1, SMC1 and FANCD2, have been implicated in the control of S-phase checkpoint. Cdc25A is the downstream effector of CHK1 and CHK2, though the underlying mechanism for control of intra S-phase checkpoint by other targets remain obscure. G2 checkpoint prevents mitotic entry solely through inhibitory phosphorylation of Cdc2/Cdk1. Several ATM targets including CHK1, CHK2, BRCA1, MDC1 and p53BP1 have been implicated in the control of G2/M

  16. The final checkpoint. Cancer as an adaptive evolutionary mechanism

    Directory of Open Access Journals (Sweden)

    Rumena Petkova

    2016-05-01

    Full Text Available The mechanisms for identification of DNA damage and repair usually manage DNA damage very efficiently. If damaged cells manage to bypass the checkpoints where the integrity of the genome is assessed and the decisions whether to proceed with the cell cycle are made, they may evade the imperative to stop dividing and to die. As a result, cancer may develop. Warding off the potential sequence-altering effects of DNA damage during the life of the individual or the existence span of the species is controlled by a set of larger checkpoints acting on a progressively increasing scale, from systematic removal of damaged cells from the proliferative pool by means of repair of DNA damage/programmed cell death through ageing to, finally, cancer. They serve different purposes and act at different levels of the life cycle, safeguarding the integrity of the genetic backup of the individual, the genetic diversity of the population, and, finally, the survival of the species and of life on Earth. In the light of the theory that cancer is the final checkpoint or the nature's manner to prevent complex organisms from living forever at the expense of genetic stagnation, the eventual failure of modern anti-cancer treatments is only to be expected. Nevertheless, the medicine of today and the near future has enough potential to slow down the progression to terminal cancer so that the life expectancy and the quality of life of cancer-affected individuals may be comparable to those of healthy aged individuals.

  17. Implication of the G2 checkpoint in the maintenance of genome integrity

    International Nuclear Information System (INIS)

    Piette, J.; Munoz, P.

    2000-01-01

    Checkpoints are surveillance mechanisms that block transitions, for instance in response to DNA damage. We summarize here here recent progress in the molecular characterization of the G 2 checkpoint which controls the entry into mitosis, and review new evidence which implicates de-regulated expression of checkpoint proteins and proteins involved in DNA damage repair in cancer development. These now exists good evidence that individuals who inherited mutations in genes involved in G 2 checkpoint and DNA damage repair are predisposed to the development of various types of cancer, their cells having a strong tendency to accumulate additional mutations. However, the occurrence of mutations of most of these genes in sporadic tumors has yet to be analysed more accurately. (authors)

  18. Participation of SRM5/CDC28, SRM8/NET1 and SRM12/HF11 genes in activation of checkpoints of Yeast Saccharomyces Cerevisiae

    International Nuclear Information System (INIS)

    Kadyshevskaya, E.Yu.; Koltovaya, N.A.

    2007-01-01

    It is known that there are about twenty checkpoint genes in yeast Saccharomyces cerevisiae. We study participation of SRM genes selected as genes affecting genetic stability and radiosensitivity. It has been shown that srm5/cdc28-srm, srm8/net1-srm, srm12/hfil-srm mutations prevent checkpoint activation by DNA damage, particularly G0/S-checkpoint (srm5, srm8), G1/S-checkpoint (srm5, srm8, srm12), S-checkpoint (srm5, srm12) and G2-checkpoint (srm5). These data indicate, at least in budding yeast, CDC28/SRM5, HF11/ADA1/SRM12 and NET1/SRM8 genes mediate cellular response induced by DNA damage including checkpoint control

  19. Efficient Incremental Checkpointing of Java Programs

    DEFF Research Database (Denmark)

    Lawall, Julia Laetitia; Muller, Gilles

    2000-01-01

    This paper investigates the optimization of language-level checkpointing of Java programs. First, we describe how to systematically associate incremental checkpoints with Java classes. While being safe, the genericness of this solution induces substantial execution overhead. Second, to solve...

  20. Network support for system initiated checkpoints

    Science.gov (United States)

    Chen, Dong; Heidelberger, Philip

    2013-01-29

    A system, method and computer program product for supporting system initiated checkpoints in parallel computing systems. The system and method generates selective control signals to perform checkpointing of system related data in presence of messaging activity associated with a user application running at the node. The checkpointing is initiated by the system such that checkpoint data of a plurality of network nodes may be obtained even in the presence of user applications running on highly parallel computers that include ongoing user messaging activity.

  1. Checkpointing for a hybrid computing node

    Science.gov (United States)

    Cher, Chen-Yong

    2016-03-08

    According to an aspect, a method for checkpointing in a hybrid computing node includes executing a task in a processing accelerator of the hybrid computing node. A checkpoint is created in a local memory of the processing accelerator. The checkpoint includes state data to restart execution of the task in the processing accelerator upon a restart operation. Execution of the task is resumed in the processing accelerator after creating the checkpoint. The state data of the checkpoint are transferred from the processing accelerator to a main processor of the hybrid computing node while the processing accelerator is executing the task.

  2. The CD47-SIRPα signaling axis as an innate immune checkpoint in cancer.

    Science.gov (United States)

    Matlung, Hanke L; Szilagyi, Katka; Barclay, Neil A; van den Berg, Timo K

    2017-03-01

    Immune checkpoint inhibitors, including those targeting CTLA-4/B7 and the PD-1/PD-L1 inhibitory pathways, are now available for clinical use in cancer patients, with other interesting checkpoint inhibitors being currently in development. Most of these have the purpose to promote adaptive T cell-mediated immunity against cancer. Here, we review another checkpoint acting to potentiate the activity of innate immune cells towards cancer. This innate immune checkpoint is composed of what has become known as the 'don't-eat me' signal CD47, which is a protein broadly expressed on normal cells and often overexpressed on cancer cells, and its counter-receptor, the myeloid inhibitory immunoreceptor SIRPα. Blocking CD47-SIRPα interactions has been shown to promote the destruction of cancer cells by phagocytes, including macrophages and neutrophils. Furthermore, there is growing evidence that targeting of the CD47-SIRPα axis may also promote antigen-presenting cell function and thereby stimulate adaptive T cell-mediated anti-cancer immunity. The development of CD47-SIRPα checkpoint inhibitors and the potential side effects that these may have are discussed. Collectively, this identifies the CD47-SIRPα axis as a promising innate immune checkpoint in cancer, and with data of the first clinical studies with CD47-SIRPα checkpoint inhibitors expected within the coming years, this is an exciting and rapidly developing field. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. An origin-deficient yeast artificial chromosome triggers a cell cycle checkpoint.

    Science.gov (United States)

    van Brabant, A J; Buchanan, C D; Charboneau, E; Fangman, W L; Brewer, B J

    2001-04-01

    Checkpoint controls coordinate entry into mitosis with the completion of DNA replication. Depletion of nucleotide precursors by treatment with the drug hydroxyurea triggers such a checkpoint response. However, it is not clear whether the signal for this hydroxyurea-induced checkpoint pathway is the presence of unreplicated DNA, or rather the persistence of single-stranded or damaged DNA. In a yeast artificial chromosome (YAC) we have engineered an approximately 170 kb region lacking efficient replication origins that allows us to explore the specific effects of unreplicated DNA on cell cycle progression. Replication of this YAC extends the length of S phase and causes cells to engage an S/M checkpoint. In the absence of Rad9 the YAC becomes unstable, undergoing deletions within the origin-free region.

  4. Blocking CHK1 Expression Induces Apoptosis and Abrogates the G2 Checkpoint Mechanism

    Directory of Open Access Journals (Sweden)

    Yan Luo

    2001-01-01

    Full Text Available Checkpoint kinase 1 (Chki is a checkpoint gene that is activated after DNA damage. It phosphorylates and inactivates the Cdc2 activating phosphatase Cdc25C. This in turn inactivates Cdc2, which leads to G2/M arrest. We report that blocking Chki expression by antisense or ribozymes in mammalian cells induces apoptosis and interferes with the G2/M arrest induced by adriamycin. The Chki inhibitor UCN-01 also blocks the G2 arrest after DNA damage and renders cells more susceptible to adriamycin. These results indicate that Chki is an essential gene for the checkpoint mechanism during normal cell proliferation as well as in the DNA damage response.

  5. Lyn tyrosine kinase promotes silencing of ATM-dependent checkpoint signaling during recovery from DNA double-strand breaks

    International Nuclear Information System (INIS)

    Fukumoto, Yasunori; Kuki, Kazumasa; Morii, Mariko; Miura, Takahito; Honda, Takuya; Ishibashi, Kenichi; Hasegawa, Hitomi; Kubota, Sho; Ide, Yudai; Yamaguchi, Noritaka; Nakayama, Yuji; Yamaguchi, Naoto

    2014-01-01

    Highlights: • Inhibition of Src family kinases decreased γ-H2AX signal. • Inhibition of Src family increased ATM-dependent phosphorylation of Chk2 and Kap1. • shRNA-mediated knockdown of Lyn increased phosphorylation of Kap1 by ATM. • Ectopic expression of Src family kinase suppressed ATM-mediated Kap1 phosphorylation. • Src is involved in upstream signaling for inactivation of ATM signaling. - Abstract: DNA damage activates the DNA damage checkpoint and the DNA repair machinery. After initial activation of DNA damage responses, cells recover to their original states through completion of DNA repair and termination of checkpoint signaling. Currently, little is known about the process by which cells recover from the DNA damage checkpoint, a process called checkpoint recovery. Here, we show that Src family kinases promote inactivation of ataxia telangiectasia mutated (ATM)-dependent checkpoint signaling during recovery from DNA double-strand breaks. Inhibition of Src activity increased ATM-dependent phosphorylation of Chk2 and Kap1. Src inhibition increased ATM signaling both in G2 phase and during asynchronous growth. shRNA knockdown of Lyn increased ATM signaling. Src-dependent nuclear tyrosine phosphorylation suppressed ATM-mediated Kap1 phosphorylation. These results suggest that Src family kinases are involved in upstream signaling that leads to inactivation of the ATM-dependent DNA damage checkpoint

  6. Non-invasive electrical brain stimulation: from acute to late-stage treatment of central nervous system damage

    Directory of Open Access Journals (Sweden)

    Petra Henrich-Noack

    2017-01-01

    Full Text Available Non-invasive brain current stimulation (NIBS is a promising and versatile tool for inducing neuroplasticity, protection and functional rehabilitation of damaged neuronal systems. It is technically simple, requires no surgery, and has significant beneficial effects. However, there are various technical approaches for NIBS which influence neuronal networks in significantly different ways. Transcranial direct current stimulation (tDCS, alternating current stimulation (ACS and repetitive transcranial magnetic stimulation (rTMS all have been applied to modulate brain activity in animal experiments under normal and pathological conditions. Also clinical trials have shown that tDCS, rTMS and ACS induce significant behavioural effects and can – depending on the parameters chosen – enhance or decrease brain excitability and influence performance and learning as well as rehabilitation and protective mechanisms. The diverse phaenomena and partially opposing effects of NIBS are not yet fully understood and mechanisms of action need to be explored further in order to select appropriate parameters for a given task, such as current type and strength, timing, distribution of current densities and electrode position. In this review, we will discuss the various parameters which need to be considered when designing a NIBS protocol and will put them into context with the envisaged applications in experimental neurobiology and medicine such as vision restoration, motor rehabilitation and cognitive enhancement.

  7. Evidence of skeletal muscle damage following electrically stimulated isometric muscle contractions in humans

    DEFF Research Database (Denmark)

    Mackey, Abigail; Bojsen-Moller, Jens; Qvortrup, Klaus

    2008-01-01

    and desmin-negative staining in a small percentage of myofibers in five and four individuals, respectively. z-Line disruption was evident at varying magnitudes in all subjects and displayed a trend toward a positive correlation (r = 0.73, P = 0.0663) with the force produced by stimulation. Increased muscle...

  8. Checkpoint inhibitors in breast cancer

    DEFF Research Database (Denmark)

    Polk, Anne; Svane, Inge-Marie; Andersson, Michael

    2018-01-01

    INTRODUCTION: An increasing number of compounds directed against immune checkpoints are currently under clinical development. In this review we summarize current research in breast cancer. MATERIAL AND METHODS: A computer-based literature search was carried out using PubMed and EMBASE; data...... reported at international meetings and clinicaltrials.gov were included as well. RESULTS: The obtained overall response rate of PD-1/PD-L1 monotherapy varied from 5 to 30% in heavily pretreated triple negative breast cancer (TNBC). The median duration of progression free survival and overall survival were...... and induce long standing anti-tumor immunity in a subgroup of breast cancer patients. However, the identification of predictive biomarkers is crucial for further development of this treatment modality....

  9. DNA-damaging agents stimulate gene expression at specific loci in Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Kenyon, C.J.; Walker, G.C.

    1988-05-01

    Operon fusions in Escherichia coli were obtained that showed increased beta-galactosidase expression in response to treatment with the DNA-damaging agent mitomycin C. These fusions were generated by using the Mud(ApR, lac) vector to insert the lactose structural genes randomly into the bacterial chromosome. Induction of beta-galactosidase in these strains, which carried fusions of lac to these din (damage-inducible) loci, was (i) triggered by UV light as well as by mitomycin C and (ii) abolished by either a recA- or a lexA- mutation. Similar characteristics of induction were observed when the lactose genes were fused to a prophage lambda promoter by using Mud(ApR, lac). These results indicate that E. coli contains a set of genes that, like prophage lambda genes, are expressed in response to DNA-damaging agents and regulated by the recA and lexA gene products. These din genes map at five bacterial loci. One din::Mud(ApR, lac) insertion results in a UV-sensitive phenotype and may be within the uvrA transcriptional unit.

  10. Caffeine stabilizes Cdc25 independently of Rad3 in S chizosaccharomyces pombe contributing to checkpoint override

    Science.gov (United States)

    Alao, John P; Sjölander, Johanna J; Baar, Juliane; Özbaki-Yagan, Nejla; Kakoschky, Bianca; Sunnerhagen, Per

    2014-01-01

    Cdc25 is required for Cdc2 dephosphorylation and is thus essential for cell cycle progression. Checkpoint activation requires dual inhibition of Cdc25 and Cdc2 in a Rad3-dependent manner. Caffeine is believed to override activation of the replication and DNA damage checkpoints by inhibiting Rad3-related proteins in both S chizosaccharomyces pombe and mammalian cells. In this study, we have investigated the impact of caffeine on Cdc25 stability, cell cycle progression and checkpoint override. Caffeine induced Cdc25 accumulation in S . pombe independently of Rad3. Caffeine delayed cell cycle progression under normal conditions but advanced mitosis in cells treated with replication inhibitors and DNA-damaging agents. In the absence of Cdc25, caffeine inhibited cell cycle progression even in the presence of hydroxyurea or phleomycin. Caffeine induces Cdc25 accumulation in S . pombe by suppressing its degradation independently of Rad3. The induction of Cdc25 accumulation was not associated with accelerated progression through mitosis, but rather with delayed progression through cytokinesis. Caffeine-induced Cdc25 accumulation appears to underlie its ability to override cell cycle checkpoints. The impact of Cdc25 accumulation on cell cycle progression is attenuated by Srk1 and Mad2. Together our findings suggest that caffeine overrides checkpoint enforcement by inducing the inappropriate nuclear localization of Cdc25. PMID:24666325

  11. DNA damage and polyploidization.

    Science.gov (United States)

    Chow, Jeremy; Poon, Randy Y C

    2010-01-01

    A growing body of evidence indicates that polyploidization triggers chromosomal instability and contributes to tumorigenesis. DNA damage is increasingly being recognized for its roles in promoting polyploidization. Although elegant mechanisms known as the DNA damage checkpoints are responsible for halting the cell cycle after DNA damage, agents that uncouple the checkpoints can induce unscheduled entry into mitosis. Likewise, defects of the checkpoints in several disorders permit mitotic entry even in the presence of DNA damage. Forcing cells with damaged DNA into mitosis causes severe chromosome segregation defects, including lagging chromosomes, chromosomal fragments and chromosomal bridges. The presence of these lesions in the cleavage plane is believed to abort cytokinesis. It is postulated that if cytokinesis failure is coupled with defects of the p53-dependent postmitotic checkpoint pathway, cells can enter S phase and become polyploids. Progress in the past several years has unraveled some of the underlying principles of these pathways and underscored the important role of DNA damage in polyploidization. Furthermore, polyploidization per se may also be an important determinant of sensitivity to DNA damage, thereby may offer an opportunity for novel therapies.

  12. Selective Cannabinoid 2 Receptor Stimulation Reduces Tubular Epithelial Cell Damage after Renal Ischemia-Reperfusion Injury.

    Science.gov (United States)

    Pressly, Jeffrey D; Mustafa, Suni M; Adibi, Ammaar H; Alghamdi, Sahar; Pandey, Pankaj; Roy, Kuldeep K; Doerksen, Robert J; Moore, Bob M; Park, Frank

    2018-02-01

    Ischemia-reperfusion injury (IRI) is a common cause of acute kidney injury (AKI), which is an increasing problem in the clinic and has been associated with elevated rates of mortality. Therapies to treat AKI are currently not available, so identification of new targets that can be modulated to ameliorate renal damage upon diagnosis of AKI is essential. In this study, a novel cannabinoid receptor 2 (CB2) agonist, SMM-295 [3'-methyl-4-(2-(thiophen-2-yl)propan-2-yl)biphenyl-2,6-diol], was designed, synthesized, and tested in vitro and in silico. Molecular docking of SMM-295 into a CB2 active-state homology model showed that SMM-295 interacts well with key amino acids to stabilize the active state. In human embryonic kidney 293 cells, SMM-295 was capable of reducing cAMP production with 66-fold selectivity for CB2 versus cannabinoid receptor 1 and dose-dependently increased mitogen-activated protein kinase and Akt phosphorylation. In vivo testing of the CB2 agonist was performed using a mouse model of bilateral IRI, which is a common model to mimic human AKI, where SMM-295 was immediately administered upon reperfusion of the kidneys after the ischemia episode. Histologic damage assessment 48 hours after reperfusion demonstrated reduced tubular damage in the presence of SMM-295. This was consistent with reduced plasma markers of renal dysfunction (i.e., creatinine and neutrophil gelatinase-associated lipocalin) in SMM-295-treated mice. Mechanistically, kidneys treated with SMM-295 were shown to have elevated activation of Akt with reduced terminal deoxynucleotidyl transferase-mediated digoxigenin-deoxyuridine nick-end labeling (TUNEL)-positive cells compared with vehicle-treated kidneys after IRI. These data suggest that selective CB2 receptor activation could be a potential therapeutic target in the treatment of AKI. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

  13. A phospho-proteomic screen identifies substrates of the checkpoint kinase Chk1

    DEFF Research Database (Denmark)

    Blasius, Melanie; Forment, Josep V; Thakkar, Neha

    2011-01-01

    BACKGROUND: The cell-cycle checkpoint kinase Chk1 is essential in mammalian cells due to its roles in controlling processes such as DNA replication, mitosis and DNA-damage responses. Despite its paramount importance, how Chk1 controls these functions remains unclear, mainly because very few Chk1...

  14. Increased sensitivity of DNA damage response-deficient cells to stimulated microgravity-induced DNA lesions.

    Directory of Open Access Journals (Sweden)

    Nan Li

    Full Text Available Microgravity is a major stress factor that astronauts have to face in space. In the past, the effects of microgravity on genomic DNA damage were studied, and it seems that the effect on genomic DNA depends on cell types and the length of exposure time to microgravity or simulated microgravity (SMG. In this study we used mouse embryonic stem (MES and mouse embryonic fibroblast (MEF cells to assess the effects of SMG on DNA lesions. To acquire the insight into potential mechanisms by which cells resist and/or adapt to SMG, we also included Rad9-deleted MES and Mdc1-deleted MEF cells in addition to wild type cells in this study. We observed significant SMG-induced DNA double strand breaks (DSBs in Rad9-/- MES and Mdc1-/- MEF cells but not in their corresponding wild type cells. A similar pattern of DNA single strand break or modifications was also observed in Rad9-/- MES. As the exposure to SMG was prolonged, Rad9-/- MES cells adapted to the SMG disturbance by reducing the induced DNA lesions. The induced DNA lesions in Rad9-/- MES were due to SMG-induced reactive oxygen species (ROS. Interestingly, Mdc1-/- MEF cells were only partially adapted to the SMG disturbance. That is, the induced DNA lesions were reduced over time, but did not return to the control level while ROS returned to a control level. In addition, ROS was only partially responsible for the induced DNA lesions in Mdc1-/- MEF cells. Taken together, these data suggest that SMG is a weak genomic DNA stress and can aggravate genomic instability in cells with DNA damage response (DDR defects.

  15. Bir1 Deletion Causes Malfunction of the Spindle Assembly Checkpoint and Apoptosis in Yeast

    International Nuclear Information System (INIS)

    Ren, Qun; Liou, Liang-Chun; Gao, Qiuqiang; Bao, Xiaoming; Zhang, Zhaojie

    2012-01-01

    Cell division in yeast is a highly regulated and well studied event. Various checkpoints are placed throughout the cell cycle to ensure faithful segregation of sister chromatids. Unexpected events, such as DNA damage or oxidative stress, cause the activation of checkpoint(s) and cell cycle arrest. Malfunction of the checkpoints may induce cell death. We previously showed that under oxidative stress, the budding yeast cohesin Mcd1, a homolog of human Rad21, was cleaved by the caspase-like protease Esp1. The cleaved Mcd1 C-terminal fragment was then translocated to mitochondria, causing apoptotic cell death. In the present study, we demonstrated that Bir1 plays an important role in spindle assembly checkpoint and cell death. Similar to H 2 O 2 treatment, deletion of BIR1 using a BIR1-degron strain caused degradation of the securin Pds1, which binds and inactivates Esp1 until metaphase-anaphase transition in a normal cell cycle. BIR1 deletion caused an increase level of ROS and mis-location of Bub1, a major protein for spindle assembly checkpoint. In wild type, Bub1 was located at the kinetochores, but was primarily in the cytoplasm in bir1 deletion strain. When BIR1 was deleted, addition of nocodazole was unable to retain the Bub1 localization on kinetochores, further suggesting that Bir1 is required to activate and maintain the spindle assembly checkpoint. Our study suggests that the BIR1 function in cell cycle regulation works in concert with its anti-apoptosis function.

  16. Peripheral benzodiazepines receptor (PBR stimulates steroidogenesis: A potential neuroprotective pathway following brain damage

    Directory of Open Access Journals (Sweden)

    George E. Barreto

    2015-04-01

    Full Text Available The effects of neuroactive steroids have been highly assessed for their significance on inflammation resolution induced by cytotoxic agents. Steroids are derived from cholesterol, and this regulatory pathway may be a target for possible protective strategies. For example, the increased expression of peripheral benzodiazepine receptor (PBR stimulates steroids production, and the action of specific ligands on PBR favors the reduction of glial activity and act as a protective mechanism. The augmented expression of PBR and steroidogenic acute regulatory protein (StAR after injury is associated with local production of steroids by glial cells. For instance, cholesterol is captured by StAR in the outer mitochondrial membrane that transfers it to PBR, which uses it as substrate for the enzyme P450scc in the inner mitochondrial membrane. Some ligands, such as 4'-Chlorodiazepam (Ro5-4864 and isoquinoline carboxamide (PK 11195, act as agonists of the PBR receptor. Previous studies indicate that Ro5-4864 reduces neuronal loss, thus implying the regulation of mitochondrial transition after a traumatic brain injury. In this work, we assess the effects of PBR ligands directly involved in neuronal cell survival and proliferation after injury, thereby activating potential downstream targets as novel therapeutic approaches.

  17. CCK1-Receptor Stimulation Protects Against Gut Mediator-Induced Lung Damage During Endotoxemia

    Directory of Open Access Journals (Sweden)

    Friederike Eisner

    2013-12-01

    Full Text Available Background/Aims: Cholecystokinin 1-receptor (CCK1-R activation by long chain fatty acid (LCFA absorption stimulates vago-vagal reflex pathways in the brain stem. The present study determines whether this reflex also activates the cholinergic anti-inflammatory pathway, a pathway known to modulate cytokine release during endotoxemia. Methods:Mesenteric lymph was obtained from wild type (WT and CCK1-R knockout (CCK1-R-/- mice intraperitoneally challenged with Lipopolysaccharid (LPS (endotoxemic lymph, EL and intestinally infused with vehicle or LCFA-enriched solution. The lymph was analyzed for TNFα, IL-6 and IL-10 concentration and administered to healthy recipient mice via jugular infusion. Alveolar wall thickness, myeloperoxidase (MPO and TUNEL positive cells were determined in lung tissue of recipient mice. Results: LCFA infusion in WT mice reduced TNFα concentration in EL by 49% compared to vehicle infusion, but had no effect in CCK1-R-/- mice. EL significantly increased the alveolar wall thickness, the number of MPO-positive and TUNEL-positive cells compared to control lymph administration. LCFA infusion in WT, but not in CCK1R-/- mice, significantly reduced these pathological effects of EL. Conclusion: During endotoxemia enteral LCFA absorption reduces TNFα release into mesenteric lymph and attenuates histomorphologic parameters of lung dysfunction. Failure to elicit this effect in CCK1R-/- mice demonstrates that anti-inflammatory properties of LCFAs are mediated through CCK1-Rs.

  18. Mistletoe (Viscum album) infestation in the Scots pine stimulates drought-dependent oxidative damage in summer.

    Science.gov (United States)

    Mutlu, Salih; Ilhan, Veli; Turkoglu, Halil Ibrahim

    2016-04-01

    This study sought to contribute to the understanding of the detrimental effect of the mistletoe (Viscum albumL.), a hemiparasitic plant, on the mortality of the Scots pine (Pinus sylvestrisL.). Fieldwork was conducted in the town of Kelkit (Gumushane province, Turkey) from April to October in 2013. Pine needles of similar ages were removed from the branches of mistletoe-infested and noninfested Scots pine plants, then transported to the laboratory and used as research materials. The effects of the mistletoe on the Scots pine during infestation were evaluated by determining the levels of water, electrolyte leakage (EL), malondialdehyde (MDA, being a product of lipid peroxidation) and reactive oxygen species (ROS) such as superoxide anion (O2 (-•)), hydrogen peroxide (H2O2) and hydroxyl radical ((•)OH). In addition, the activities of antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT) and peroxidase (POX) were measured in the same samples. The highest level of drought stress was found in summer (especially in August) as a result of the lowest water content in the soil and the highest average temperature occurring in these months. The drought stress induced by mistletoe infestation caused a regular decrease in water content, while it increased the levels of EL, MDA and ROS (H2O2, O2 (-•)and(•)OH). The infestation also stimulated the activities of CAT and POX, with the exception of SOD. On the other hand, in August, when the drought conditions were the harshest, the levels of EL and MDA, which are two of the most important indicator parameters for oxidative stress, as well as the levels of H2O2and(•)OH, which are two of the ROS leading to oxidative stress, reached the highest values in both infested and noninfested needles, whereas the O2 (-•)level decreased. For the same period and needles, CAT activity increased, while SOD activity decreased. Peroxidase activity, however, did not exhibit a significant change. Our findings indicate

  19. Asynchronous Checkpoint Migration with MRNet in the Scalable Checkpoint / Restart Library

    Energy Technology Data Exchange (ETDEWEB)

    Mohror, K; Moody, A; de Supinski, B R

    2012-03-20

    Applications running on today's supercomputers tolerate failures by periodically saving their state in checkpoint files on stable storage, such as a parallel file system. Although this approach is simple, the overhead of writing the checkpoints can be prohibitive, especially for large-scale jobs. In this paper, we present initial results of an enhancement to our Scalable Checkpoint/Restart Library (SCR). We employ MRNet, a tree-based overlay network library, to transfer checkpoints from the compute nodes to the parallel file system asynchronously. This enhancement increases application efficiency by removing the need for an application to block while checkpoints are transferred to the parallel file system. We show that the integration of SCR with MRNet can reduce the time spent in I/O operations by as much as 15x. However, our experiments exposed new scalability issues with our initial implementation. We discuss the sources of the scalability problems and our plans to address them.

  20. Checkpoint Kinase 1 Expression Predicts Poor Prognosis in Nigerian Breast Cancer Patients.

    Science.gov (United States)

    Ebili, Henry Okuchukwu; Iyawe, Victoria O; Adeleke, Kikelomo Rachel; Salami, Babatunde Abayomi; Banjo, Adekunbiola Aina; Nolan, Chris; Rakha, Emad; Ellis, Ian; Green, Andrew; Agboola, Ayodeji Olayinka Johnson

    2018-02-01

    Checkpoint kinase 1 (CHEK1), a DNA damage sensor and cell death pathway stimulator, is regarded as an oncogene in tumours, where its activities are considered essential for tumourigenesis and the survival of cancer cells treated with chemotherapy and radiotherapy. In breast cancer, CHEK1 expression has been associated with an aggressive tumour phenotype, the triple-negative breast cancer subtype, an aberrant response to tamoxifen, and poor prognosis. However, the relevance of CHEK1 expression has, hitherto, not been investigated in an indigenous African population. We therefore aimed to investigate the clinicopathological, biological, and prognostic significance of CHEK1 expression in a cohort of Nigerian breast cancer cases. Tissue microarrays of 207 Nigerian breast cancer cases were tested for CHEK1 expression using immunohistochemistry. The clinicopathological, molecular, and prognostic characteristics of CHEK1-positive tumours were determined using the Chi-squared test and Kaplan-Meier and Cox regression analyses in SPSS Version 16. Nuclear expression of CHEK1 was present in 61% of breast tumours and was associated with tumour size, triple-negative cancer, basal-like phenotype, the epithelial-mesenchymal transition, p53 over-expression, DNA homologous repair pathway dysfunction, and poor prognosis. The rate expression of CHEK1 is high in Nigerian breast cancer cases and is associated with an aggressive phenotype and poor prognosis.

  1. [Effects of electric stimulation at the cerebellar fastigial nucleus on astrocytes in the hippocampus of neonatal rats with hypoxic-ischemic brain damage].

    Science.gov (United States)

    Li, Xiao-Li; Jia, Tian-Ming; Luan, Bin; Liu, Tao; Yuan, Yan

    2011-04-01

    To study the effects of electric stimulation at the cerebellar fastigial nucleus on astrocytes in the hippocampus of neonatal rats with hypoxic-ischemic brain damage (HIBD) and the possible mechanism. One hundred and eighty 7-day-old neonatal Sprague-Dawley rats were randomly divided into three groups: sham-operation (control group) and HIBD with and without electric stimulation (n=60 each). The HIBD model of neonatal rats was prepared by the Rice-Vennucci method. Electric stimulation at the cerebellar fastigial nucleus was given 24 hrs after the operation in the electric stimulation group once daily and lasted for 30 minutes each time. The other two groups were not subjected to electric stimulation but captured to fix in corresponding periods. Rats were sacrificed 3, 7, 14 and 21 days after stimulations to observe the glial fibrillary acidic protein (GFAP) expression by immunohistochemisty and the ultrastructural changes of astrocytes in the hippocampus under an electron microscope. Immunohistochemical analysis showed the expression of GFAP in the HIBD groups with and without electric stimulation increased significantly compared with the control group on day 3, reached the peak on day 7, and the increased expression remained till to day 21. The GFAP expression in the electric stimulation group was significantly lower than that in the untreated HIBD group at all time points. Under the electron microscope, the astrocytes in the untreated HIBD group were swollen and the amount of organelles was reduced, while the swelling of astrocytes was alleviated and the organelles remained in integrity in the electric stimulation group. The electric stimulation at the cerebellar fastigial nucleus can inhibit the excessive proliferation of astrocytes and relieve the structural damage of astrocytes in neonatal rats following HIBD.

  2. DNA damage in the oocytes SACs

    Czech Academy of Sciences Publication Activity Database

    Macůrek, Libor

    2016-01-01

    Roč. 15, č. 4 (2016), s. 491-492 ISSN 1538-4101 Institutional support: RVO:68378050 Keywords : DNA damage response * oocyte * meiosis * checkpoint Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.530, year: 2016

  3. Molecular Mechanisms of DNA Replication Checkpoint Activation

    Directory of Open Access Journals (Sweden)

    Bénédicte Recolin

    2014-03-01

    Full Text Available The major challenge of the cell cycle is to deliver an intact, and fully duplicated, genetic material to the daughter cells. To this end, progression of DNA synthesis is monitored by a feedback mechanism known as replication checkpoint that is untimely linked to DNA replication. This signaling pathway ensures coordination of DNA synthesis with cell cycle progression. Failure to activate this checkpoint in response to perturbation of DNA synthesis (replication stress results in forced cell division leading to chromosome fragmentation, aneuploidy, and genomic instability. In this review, we will describe current knowledge of the molecular determinants of the DNA replication checkpoint in eukaryotic cells and discuss a model of activation of this signaling pathway crucial for maintenance of genomic stability.

  4. Process of radiation damage in BaFX:Eu2+ (X=Br, I) photo-stimulable phosphor for imaging plate (IP)

    International Nuclear Information System (INIS)

    Kato, Munetaka; Takahashi, Kenji

    2005-01-01

    Imaging Plate (IP) is a 2-dimensional radiation detector utilizing photo-stimulable phosphors. Photo-stimulated luminescence (PSL) is a kind of process in radiation-induced reactions and IP can be repeatedly performed. The mechanism of PSL is reviewed and the radiation damage, which is a phenomenon that the luminescence goes down under X-ray irradiation, is also referred to. The relation between the composition of BaFX: Eu 2+ (X=Br, I), which is widely used in IPs, and the degree of radiation damage is investigated. The degree of radiation damage is smaller in the mixed crystal system of Br and I such as BaFBr 0.85 I 0.15 :Eu 2+ than in BaFBr: Eu 2+ and BaFI: Eu 2+ . This is explained by the characteristics of luminescence centers and F centers under X-ray irradiation. (author)

  5. Cells bearing chromosome aberrations lacking one telomere are selectively blocked at the G2/M checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez, Pilar [Unitat de Biologia Cel.lular, Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Barquinero, Joan Francesc [Unitat d' Antropologia Biologica, Departament de Biologia Animal, Biologia Vegetal i Ecologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Duran, Assumpta [Unitat de Biologia Cel.lular, Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Caballin, Maria Rosa [Unitat d' Antropologia Biologica, Departament de Biologia Animal, Biologia Vegetal i Ecologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain); Ribas, Montserrat [Servei de Radiofisica i Radioproteccio de l' Hospital de la Santa Creu i Sant Pau, 08025 Barcelona (Spain); Barrios, Leonardo, E-mail: Lleonard.Barrios@uab.cat [Unitat de Biologia Cel.lular, Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Bellaterra (Spain)

    2009-11-02

    Cell cycle checkpoints are part of the cellular mechanisms to maintain genomic integrity. After ionizing radiation exposure, the cells can show delay or arrest in their progression through the cell cycle, as well as an activation of the DNA repair machinery in order to reduce the damage. The G2/M checkpoint prevents G2 cells entering mitosis until the DNA damage has been reduced. The present study evaluates which G0 radiation-induced chromosome aberrations are negatively selected in the G2/M checkpoint. For this purpose, peripheral blood samples were irradiated at 1 and 3 Gy of {gamma}-rays, and lymphocytes were cultured for 48 h. Calyculin-A and Colcemid were used to analyze, in the same slide, cells in G2 and M. Chromosome spreads were consecutively analyzed by solid stain, pancentromeric and pantelomeric FISH and mFISH. The results show that the frequency of incomplete chromosome elements, those lacking a telomeric signal at one end, decreases abruptly from G2 to M. This indicates that cells with incomplete chromosome elements can progress from G0 to G2, but at the G2/M checkpoint suffer a strong negative selection.

  6. Cells bearing chromosome aberrations lacking one telomere are selectively blocked at the G2/M checkpoint

    International Nuclear Information System (INIS)

    Rodriguez, Pilar; Barquinero, Joan Francesc; Duran, Assumpta; Caballin, Maria Rosa; Ribas, Montserrat; Barrios, Leonardo

    2009-01-01

    Cell cycle checkpoints are part of the cellular mechanisms to maintain genomic integrity. After ionizing radiation exposure, the cells can show delay or arrest in their progression through the cell cycle, as well as an activation of the DNA repair machinery in order to reduce the damage. The G2/M checkpoint prevents G2 cells entering mitosis until the DNA damage has been reduced. The present study evaluates which G0 radiation-induced chromosome aberrations are negatively selected in the G2/M checkpoint. For this purpose, peripheral blood samples were irradiated at 1 and 3 Gy of γ-rays, and lymphocytes were cultured for 48 h. Calyculin-A and Colcemid were used to analyze, in the same slide, cells in G2 and M. Chromosome spreads were consecutively analyzed by solid stain, pancentromeric and pantelomeric FISH and mFISH. The results show that the frequency of incomplete chromosome elements, those lacking a telomeric signal at one end, decreases abruptly from G2 to M. This indicates that cells with incomplete chromosome elements can progress from G0 to G2, but at the G2/M checkpoint suffer a strong negative selection.

  7. Differential impact of diverse anticancer chemotherapeutics on the Cdc25A-degradation checkpoint pathway

    International Nuclear Information System (INIS)

    Agner, Jeppe; Falck, Jacob; Lukas, Jiri; Bartek, Jiri

    2005-01-01

    When exposed to DNA-damaging insults such as ionizing radiation (IR) or ultraviolet light (UV), mammalian cells activate checkpoint pathways to halt cell cycle progression or induce cell death. Here we examined the ability of five commonly used anticancer drugs with different mechanisms of action to activate the Chk1/Chk2-Cdc25A-CDK2/cyclin E cell cycle checkpoint pathway, previously shown to be induced by IR or UV. Whereas exposure of human cells to topoisomerase inhibitors camptothecin, etoposide, or adriamycin resulted in rapid (within 1 h) activation of the pathway including degradation of the Cdc25A phosphatase and inhibition of cyclin E/CDK2 kinase activity, taxol failed to activate this checkpoint even after a prolonged treatment. Unexpectedly, although the alkylating agent cisplatin also induced degradation of Cdc25A (albeit delayed, after 8-12 h), cyclin E/CDK2 activity was elevated and DNA synthesis continued, a phenomena that correlated with increased E2F1 protein levels and consequently enhanced expression of cyclin E. These results reveal a differential impact of various classes of anticancer chemotherapeutics on the Cdc25A-degradation pathway, and indicate that the kinetics of checkpoint induction, and the relative balance of key components within the DNA damage response network may dictate whether the treated cells arrest their cell cycle progression

  8. Transparent checkpointing and process migration in a distributed system

    OpenAIRE

    2004-01-01

    A distributed system for creating a checkpoint for a plurality of processes running on the distributed system. The distributed system includes a plurality of compute nodes with an operating system executing on each compute node. A checkpoint library resides at the user level on each of the compute nodes, and the checkpoint library is transparent to the operating system residing on the same compute node and to the other compute nodes. Each checkpoint library uses a windowed messaging logging p...

  9. Multiple functions of the S-phase checkpoint mediator.

    Science.gov (United States)

    Tanaka, Katsunori

    2010-01-01

    There is mounting evidence that replication defects are the major source of spontaneous genomic instability in cells, and that S-phase checkpoints are the principal defense against such instability. The S-phase checkpoint mediator protein Mrc1/Claspin mediates the checkpoint response to replication stress by facilitating phosphorylation of effector kinase by a sensor kinase. In this review, the multiple functions and the regulation of the S-phase checkpoint mediator are discussed.

  10. A novel ATM-dependent checkpoint defect distinct from loss of function mutation promotes genomic instability in melanoma.

    Science.gov (United States)

    Spoerri, Loredana; Brooks, Kelly; Chia, KeeMing; Grossman, Gavriel; Ellis, Jonathan J; Dahmer-Heath, Mareike; Škalamera, Dubravka; Pavey, Sandra; Burmeister, Bryan; Gabrielli, Brian

    2016-05-01

    Melanomas have high levels of genomic instability that can contribute to poor disease prognosis. Here, we report a novel defect of the ATM-dependent cell cycle checkpoint in melanoma cell lines that promotes genomic instability. In defective cells, ATM signalling to CHK2 is intact, but the cells are unable to maintain the cell cycle arrest due to elevated PLK1 driving recovery from the arrest. Reducing PLK1 activity recovered the ATM-dependent checkpoint arrest, and over-expressing PLK1 was sufficient to overcome the checkpoint arrest and increase genomic instability. Loss of the ATM-dependent checkpoint did not affect sensitivity to ionizing radiation demonstrating that this defect is distinct from ATM loss of function mutations. The checkpoint defective melanoma cell lines over-express PLK1, and a significant proportion of melanomas have high levels of PLK1 over-expression suggesting this defect is a common feature of melanomas. The inability of ATM to impose a cell cycle arrest in response to DNA damage increases genomic instability. This work also suggests that the ATM-dependent checkpoint arrest is likely to be defective in a higher proportion of cancers than previously expected. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. Checkpointing and Recovery in Distributed and Database Systems

    Science.gov (United States)

    Wu, Jiang

    2011-01-01

    A transaction-consistent global checkpoint of a database records a state of the database which reflects the effect of only completed transactions and not the results of any partially executed transactions. This thesis establishes the necessary and sufficient conditions for a checkpoint of a data item (or the checkpoints of a set of data items) to…

  12. Immune mediated neuropathy following checkpoint immunotherapy.

    Science.gov (United States)

    Gu, Yufan; Menzies, Alexander M; Long, Georgina V; Fernando, S L; Herkes, G

    2017-11-01

    Checkpoint immunotherapy has revolutionised cancer therapy and is now standard treatment for many malignancies including metastatic melanoma. Acute inflammatory neuropathies, often labelled as Guillain-Barre syndrome, are an uncommon but potentially severe complication of checkpoint immunotherapy with individual cases described but never characterised as a group. We describe a case of acute sensorimotor and autonomic neuropathy following a single dose of combination ipilimumab and nivolumab for metastatic melanoma. A literature search was performed, identifying 14 other cases of acute neuropathy following checkpoint immunotherapy, with the clinical, electrophysiological and laboratory features summarised. Most cases described an acute sensorimotor neuropathy (92%) with hyporeflexia (92%) that could occur from induction up till many weeks after the final dose of therapy. In contrast to Guillain-Barre syndrome, the cerebrospinal fluid (CSF) analysis often shows a lymphocytic picture (50%) and the electrophysiology showed an axonal pattern (55%). Treatment was variable and often in combination. 11 cases received steroid therapy with only 1 death within this group, whereas of the 4 patients who did not receive steroid therapy there were 3 deaths. In conclusion checkpoint immunotherapy - induced acute neuropathies are distinct from and progress differently to Guillain-Barre syndrome. As with other immunotherapy related adverse events corticosteroid therapy should be initiated in addition to usual therapy. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Dpb11/TopBP1 plays distinct roles in DNA replication, checkpoint response and homologous recombination

    DEFF Research Database (Denmark)

    Germann, Susanne Manuela; Østergaard, Vibe Hallundbæk; Haas, Caroline

    2011-01-01

    DPB11/TopBP1 is an essential evolutionarily conserved gene involved in initiation of DNA replication and checkpoint signaling. Here, we show that Saccharomyces cerevisiae Dpb11 forms nuclear foci that localize to sites of DNA damage in G1, S and G2 phase, a recruitment that is conserved for its...... and Tel1, and of the checkpoint mediator Rad9. In a site-directed mutagenesis screen, we identify a separation-of-function mutant, dpb11-PF, that is sensitive to DSB-inducing agents yet remains proficient for DNA replication and the S-phase checkpoint at the permissive temperature. The dpb11-PF mutant...... homologue TopBP1 in Gallus gallus. Damage-induced Dpb11 foci are distinct from Sld3 replication initiation foci. Further, Dpb11 foci are dependent on the checkpoint proteins Mec3 (9-1-1 complex) and Rad24, and require the C-terminal domain of Dpb11. Dpb11 foci are independent of the checkpoint kinases Mec1...

  14. Detailed Modeling and Evaluation of a Scalable Multilevel Checkpointing System

    Energy Technology Data Exchange (ETDEWEB)

    Mohror, Kathryn [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Moody, Adam [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Bronevetsky, Greg [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); de Supinski, Bronis R. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2014-09-01

    High-performance computing (HPC) systems are growing more powerful by utilizing more components. As the system mean time before failure correspondingly drops, applications must checkpoint frequently to make progress. But, at scale, the cost of checkpointing becomes prohibitive. A solution to this problem is multilevel checkpointing, which employs multiple types of checkpoints in a single run. Moreover, lightweight checkpoints can handle the most common failure modes, while more expensive checkpoints can handle severe failures. We designed a multilevel checkpointing library, the Scalable Checkpoint/Restart (SCR) library, that writes lightweight checkpoints to node-local storage in addition to the parallel file system. We present probabilistic Markov models of SCR's performance. We show that on future large-scale systems, SCR can lead to a gain in machine efficiency of up to 35 percent, and reduce the load on the parallel file system by a factor of two. In addition, we predict that checkpoint scavenging, or only writing checkpoints to the parallel file system on application termination, can reduce the load on the parallel file system by 20 × on today's systems and still maintain high application efficiency.

  15. Wip1 phosphatase is associated with chromatin and dephosphorylates gammaH2AX to promote checkpoint inhibition

    Czech Academy of Sciences Publication Activity Database

    Macůrek, Libor; Lindqvist, A.; Voets, O.; Kool, J.; Vos, H.R.; Medema, R.H.

    2010-01-01

    Roč. 29, č. 15 (2010), s. 2281-2291 ISSN 0950-9232 R&D Projects: GA ČR GPP305/10/P420 Institutional research plan: CEZ:AV0Z50520514 Keywords : DNA damage * checkpoint * phosphatase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.414, year: 2010

  16. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis.

    Science.gov (United States)

    Pushpavalli, Sreerangam N C V L; Sarkar, Arpita; Ramaiah, M Janaki; Chowdhury, Debabani Roy; Bhadra, Utpal; Pal-Bhadra, Manika

    2013-01-24

    In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk) is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof¹/+; mnkp⁶/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using Drosophila as model system and carry out the interaction of MOF

  17. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis

    Directory of Open Access Journals (Sweden)

    Pushpavalli Sreerangam NCVL

    2013-01-01

    Full Text Available Abstract Background In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Results Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof1/+; mnkp6/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. Conclusion mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using

  18. Extending the Binomial Checkpointing Technique for Resilience

    Energy Technology Data Exchange (ETDEWEB)

    Walther, Andrea; Narayanan, Sri Hari Krishna

    2016-10-10

    In terms of computing time, adjoint methods offer a very attractive alternative to compute gradient information, re- quired, e.g., for optimization purposes. However, together with this very favorable temporal complexity result comes a memory requirement that is in essence proportional with the operation count of the underlying function, e.g., if algo- rithmic differentiation is used to provide the adjoints. For this reason, checkpointing approaches in many variants have become popular. This paper analyzes an extension of the so-called binomial approach to cover also possible failures of the computing systems. Such a measure of precaution is of special interest for massive parallel simulations and adjoint calculations where the mean time between failure of the large scale computing system is smaller than the time needed to complete the calculation of the adjoint information. We de- scribe the extensions of standard checkpointing approaches required for such resilience, provide a corresponding imple- mentation and discuss numerical results.

  19. Immune checkpoint inhibitors for metastatic bladder cancer.

    Science.gov (United States)

    Massari, Francesco; Di Nunno, Vincenzo; Cubelli, Marta; Santoni, Matteo; Fiorentino, Michelangelo; Montironi, Rodolfo; Cheng, Liang; Lopez-Beltran, Anto; Battelli, Nicola; Ardizzoni, Andrea

    2018-03-01

    Chemotherapy has represented the standard therapy for unresectable or metastatic urothelial carcinoma for more than 20 years. The growing knowledge of the interaction between tumour and immune system has led to the advent of new classes of drugs, the immune-checkpoints inhibitors, which are intended to change the current scenario. To date, immunotherapy is able to improve the overall responses and survival. Moreover, thanks to its safety profile immune-checkpoint inhibitors could be proposed also to patients unfit for standard chemotherapy. No doubts that these agents have started a revolution expected for years, but despite this encouraging results it appears clear that not all subjects respond to these agents and requiring the development of reliable predictive response factors able to isolate patients who can more benefit from these treatments as well as new strategies aimed to improve immunotherapy clinical outcome. In this review we describe the active or ongoing clinical trials involving Programmed Death Ligand 1 (PD-L1), Programmed Death receptor 1 (PD-1) and Cytotoxic-T Lymphocyte Antigen 4 (CTLA 4) inhibitors in urothelial carcinoma focusing our attention on the developing new immune-agents and combination strategies with immune-checkpoint inhibitors. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Tyrosine 370 phosphorylation of ATM positively regulates DNA damage response

    Science.gov (United States)

    Lee, Hong-Jen; Lan, Li; Peng, Guang; Chang, Wei-Chao; Hsu, Ming-Chuan; Wang, Ying-Nai; Cheng, Chien-Chia; Wei, Leizhen; Nakajima, Satoshi; Chang, Shih-Shin; Liao, Hsin-Wei; Chen, Chung-Hsuan; Lavin, Martin; Ang, K Kian; Lin, Shiaw-Yih; Hung, Mien-Chie

    2015-01-01

    Ataxia telangiectasia mutated (ATM) mediates DNA damage response by controling irradiation-induced foci formation, cell cycle checkpoint, and apoptosis. However, how upstream signaling regulates ATM is not completely understood. Here, we show that upon irradiation stimulation, ATM associates with and is phosphorylated by epidermal growth factor receptor (EGFR) at Tyr370 (Y370) at the site of DNA double-strand breaks. Depletion of endogenous EGFR impairs ATM-mediated foci formation, homologous recombination, and DNA repair. Moreover, pretreatment with an EGFR kinase inhibitor, gefitinib, blocks EGFR and ATM association, hinders CHK2 activation and subsequent foci formation, and increases radiosensitivity. Thus, we reveal a critical mechanism by which EGFR directly regulates ATM activation in DNA damage response, and our results suggest that the status of ATM Y370 phosphorylation has the potential to serve as a biomarker to stratify patients for either radiotherapy alone or in combination with EGFR inhibition. PMID:25601159

  1. CDC28, NETI, and HFII are required for checkpoints in Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Koltovaya, N.A.; Kadyshevskaya, E.Yu.; Roshina, M.P.; Devin, A.B.

    2009-01-01

    The involvement of SRM genes selected as genes affecting genetic stability and radiosensitivity in a cell cycle arrest under the action of damaging agents was studied. It was shown that the srm5/cdc28-srm, srm8/netI-srm, and srmI2/hfiI-srm mutations prevent checkpoint activation by DNA damage, particularly the G 0 /S (srm5, srm8), G 1 /S (srm5, srm8, srm12), S (srm8, srm12) and S/G 2 (srm5) checkpoints. It seems that in budding yeast the CDC28, HFII/ADAI, and NETI genes mediate cellular response induced by DNA damage with checkpoint control. The well-known checkpoint-genes RAD9, RAD17, RAD24, and RAD53, and the genes CDC28, and NETI have been found to belong to one epistasis group named RAD9-group as regards cell sensitivity to γ radiation. An analysis of the radiosensitivity of double mutants has revealed that the mutation cdc-28-srm is hypostatic to each of mutations rad9Δ, and rad24Δ, and additive to rad17Δ. The mutation netI-srm is hypostatic to the mutations rad9Δ but additive to rad17Δ, rad24Δ, and rad53. The mutation hfiI-srm has an additive effect in compound with the mutations rad24Δ and rad9Δ. So, investigations of epistatic interactions have demonstrated a branched RAD9-dependent pathway. The analyzed genes can also participate in a minor mechanism involved in determining cell radiation sensitivity independently of the mentioned RAD9-dependent pathway

  2. Translational Control Protein 80 Stimulates IRES-Mediated Translation of p53 mRNA in Response to DNA Damage

    Directory of Open Access Journals (Sweden)

    Marie-Jo Halaby

    2015-01-01

    Full Text Available Synthesis of the p53 tumor suppressor increases following DNA damage. This increase and subsequent activation of p53 are essential for the protection of normal cells against tumorigenesis. We previously discovered an internal ribosome entry site (IRES that is located at the 5′-untranslated region (UTR of p53 mRNA and found that the IRES activity increases following DNA damage. However, the mechanism underlying IRES-mediated p53 translation in response to DNA damage is still poorly understood. In this study, we discovered that translational control protein 80 (TCP80 has increased binding to the p53 mRNA in vivo following DNA damage. Overexpression of TCP80 also leads to increased p53 IRES activity in response to DNA damage. TCP80 has increased association with RNA helicase A (RHA following DNA damage and overexpression of TCP80, along with RHA, leads to enhanced expression of p53. Moreover, we found that MCF-7 breast cancer cells with decreased expression of TCP80 and RHA exhibit defective p53 induction following DNA damage and diminished expression of its downstream target PUMA, a proapoptotic protein. Taken together, our discovery of the function of TCP80 and RHA in regulating p53 IRES and p53 induction following DNA damage provides a better understanding of the mechanisms that regulate IRES-mediated p53 translation in response to genotoxic stress.

  3. The Milieu of Damaged Alveolar Epithelial Type 2 Cells Stimulates Alveolar Wound Repair by Endogenous and Exogenous Progenitors

    Science.gov (United States)

    Buckley, Susan; Shi, Wei; Carraro, Gianni; Sedrakyan, Sargis; Da Sacco, Stefano; Driscoll, Barbara A.; Perin, Laura; De Filippo, Roger E.

    2011-01-01

    Alveolar epithelial integrity is dependent upon the alveolar milieu, yet the milieu of the damaged alveolar epithelial cell type 2 (AEC2) has been little studied. Characterization of its components may offer the potential for ex vivo manipulation of stem cells to optimize their therapeutic potential. We examined the cytokine profile of AEC2 damage milieu, hypothesizing that it would promote endogenous epithelial repair while recruiting cells from other locations and instructing their engraftment and differentiation. Bronchoalveolar lavage and lung extract from hyperoxic rats represented AEC2 in vivo damage milieu, and medium from a scratch-damaged AEC2 monolayer represented in vitro damage. CINC-2 and ICAM, the major cytokines detected by proteomic cytokine array in AEC2 damage milieu, were chemoattractive to normoxic AECs and expedited in vitro wound healing, which was blocked by their respective neutralizing antibodies. The AEC2 damage milieu was also chemotactic for exogenous uncommitted human amniotic fluid stem cells (hAFSCs), increasing migration greater than 20-fold. hAFSCs attached within an in vitro AEC2 wound and expedited wound repair by contributing cytokines migration inhibitory factor and plasminogen activator inhibitor 1 to the AEC2 damage milieu, which promoted wound healing. The AEC2 damage milieu also promoted differentiation of a subpopulation of hAFSCs to express SPC, TTF-1, and ABCA3, phenotypic markers of distal alveolar epithelium. Thus, the microenvironment created by AEC2 damage not only promotes autocrine repair but also can attract uncommitted stem cells, which further augment healing through cytokine secretion and differentiation. PMID:21700959

  4. The relationship between retinal damage and current intensity in a pre-clinical suprachoroidal-transretinal stimulation model using a laser-formed microporous electrode

    Science.gov (United States)

    Kanda, Hiroyuki; Nakano, Yukari; Terasawa, Yasuo; Morimoto, Takeshi; Fujikado, Takashi

    2017-10-01

    Objective. Suprachoroidal-transretinal stimulation (STS) is a stimulation method for retinal prostheses. For STS-type retinal prostheses, we developed a new type of stimulating electrode called a femtosecond laser-induced porous electrode (FLiP electrode). To verify the safety of the FLiP electrode for STS, we investigated the characteristics of STS-induced retinal injury. Approach. Sixteen eyes of pigmented rabbits were studied in this in vivo study. For each examined eye, we implanted a single-channel FLiP electrode (diameter, 0.5 mm height, 0.3 mm geometric surface area, 0.43 mm2) in a scleral pocket created at the posterior pole of the eye. A return electrode (diameter, 0.5 mm length, 3 mm) was inserted into the vitreous cavity. The eyes were divided into five groups, and each group was stimulated with a different current intensity. The stimulus intensities and the number of eyes in each group were as follows: 1.0 mA (n  =  2), 1.5 mA (n  =  3), 2.0 mA (n  =  3), 2.5 mA (n  =  4), and 3.0 mA (n  =  2). Continuous biphasic pulses (0.5 ms/phase) were applied under general anesthesia at a frequency of 20 Hz for 48 h. Fundus photography, fluorescein angiography (FA), and optical coherence tomography were performed before and after applying the electrical stimulation to evaluate the retinal injury. Main results. The 1.0 mA and 1.5 mA groups showed little or no retinal damage. Fluorescent dye leakage in FA and punctate pigmentation in the fundus were observed around the stimulation site with stimulation of 2.0 mA (1/3), 2.5 mA (1/4), and 3.0 mA (2/2). Significance. Our findings indicate that the threshold current for inducing retinal damage is greater than that for eliciting electrical phosphenes (<1 mA) with STS observed in human trials. Therefore, STS by the FLiP electrode is a safe and feasible stimulation method for retinal prostheses as long as it is used with these pulse parameters.

  5. Advances of Immune Checkpoint Inhibitors in Tumor Immunotherapy

    Science.gov (United States)

    Guo, Qiao

    2018-01-01

    Immune checkpoints are cell surface molecules that can fine-tune the immune responses, they are crucial for modulating the duration and amplitude of immune reactions while maintaining self-tolerance in order to minimize autoimmune responses. Numerous studies have demonstrated that tumors cells can directly express immune-checkpoint molecules, or induce many inhibitory molecules expression in the tumor microenvironment to inhibit the anti-tumor immunity. Releasing these brakes has emerged as an exciting strategy to cure cancer. In the past few years, clinical trials with therapeutic antibodies targeting to the checkpoint molecules CTLA-4 and PD-1 have rekindled the hope for cancer immunotherapy. In contrast to the conventional treatment, checkpoint inhibitors induce broad and durable antitumor responses. In the future, treatment may involve combination therapy to target different checkpoint molecules and stages of the adaptive immune responses. In this review, we summarized the recent advances of the study and development of other checkpoint molecules in tumor immunotherapy.

  6. Template based parallel checkpointing in a massively parallel computer system

    Science.gov (United States)

    Archer, Charles Jens [Rochester, MN; Inglett, Todd Alan [Rochester, MN

    2009-01-13

    A method and apparatus for a template based parallel checkpoint save for a massively parallel super computer system using a parallel variation of the rsync protocol, and network broadcast. In preferred embodiments, the checkpoint data for each node is compared to a template checkpoint file that resides in the storage and that was previously produced. Embodiments herein greatly decrease the amount of data that must be transmitted and stored for faster checkpointing and increased efficiency of the computer system. Embodiments are directed to a parallel computer system with nodes arranged in a cluster with a high speed interconnect that can perform broadcast communication. The checkpoint contains a set of actual small data blocks with their corresponding checksums from all nodes in the system. The data blocks may be compressed using conventional non-lossy data compression algorithms to further reduce the overall checkpoint size.

  7. Enhancing Immune Checkpoint Inhibitor Therapy in Kidney Cancer

    Science.gov (United States)

    2017-10-01

    AWARD NUMBER: W81XWH-15-1-0141 TITLE: Enhancing Immune Checkpoint Inhibitor therapy in Kidney Cancer PRINCIPAL INVESTIGATOR: Hans-Joerg Hammers...SUBTITLE Enhancing Immune Checkpoint Inhibitor therapy in Kidney Cancer 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH- 15-1-0141 5c. PROGRAM ELEMENT NUMBER...immune checkpoint inhibition in kidney cancer . The work is designed to test different strategies to induce or enhance the abscopal in a kidney cancer

  8. Hypoxia‐induced alterations of G2 checkpoint regulators

    OpenAIRE

    Hasvold, Grete; Lund-Andersen, Christin; Lando, Malin; Patzke, Sebastian; Hauge, Sissel; Suo, ZhenHe; Lyng, Heidi; Syljuåsen, Randi G.

    2016-01-01

    Hypoxia promotes an aggressive tumor phenotype with increased genomic instability, partially due to downregulation of DNA repair pathways. However, genome stability is also surveilled by cell cycle checkpoints. An important issue is therefore whether hypoxia also can influence the DNA damage‐induced cell cycle checkpoints. Here, we show that hypoxia (24 h 0.2% O2) alters the expression of several G2 checkpoint regulators, as examined by microarray gene expression analysis and immunoblotting o...

  9. Checkpoint independence of most DNA replication origins in fission yeast

    OpenAIRE

    Mickle, Katie L; Ramanathan, Sunita; Rosebrock, Adam; Oliva, Anna; Chaudari, Amna; Yompakdee, Chulee; Scott, Donna; Leatherwood, Janet; Huberman, Joel A

    2007-01-01

    Abstract Background In budding yeast, the replication checkpoint slows progress through S phase by inhibiting replication origin firing. In mammals, the replication checkpoint inhibits both origin firing and replication fork movement. To find out which strategy is employed in the fission yeast, Schizosaccharomyces pombe, we used microarrays to investigate the use of origins by wild-type and checkpoint-mutant strains in the presence of hydroxyurea (HU), which limits the pool of deoxyribonucleo...

  10. The impact of neuromuscular electrical stimulation on recovery after intensive, muscle damaging, maximal speed training in professional team sports players.

    Science.gov (United States)

    Taylor, Tom; West, Daniel J; Howatson, Glyn; Jones, Chris; Bracken, Richard M; Love, Thomas D; Cook, Christian J; Swift, Eamon; Baker, Julien S; Kilduff, Liam P

    2015-05-01

    During congested fixture periods in team sports, limited recovery time and increased travel hinder the implementation of many recovery strategies; thus alternative methods are required. We examined the impact of a neuromuscular electrical stimulation device on 24-h recovery from an intensive training session in professional players. Twenty-eight professional rugby and football academy players completed this randomised and counter-balanced study, on 2 occasions, separated by 7 days. After baseline perceived soreness, blood (lactate and creatine kinase) and saliva (testosterone and cortisol) samples were collected, players completed a standardised warm-up and baseline countermovement jumps (jump height). Players then completed 60 m × 50 m maximal sprints, with 5 min recovery between efforts. After completing the sprint session, players wore a neuromuscular electrical stimulation device or remained in normal attire (CON) for 8 h. All measures were repeated immediately, 2 and 24-h post-sprint. Player jump height was reduced from baseline at all time points under both conditions; however, at 24-h neuromuscular electrical stimulation was significantly more recovered (mean±SD; neuromuscular electrical stimulation -3.2±3.2 vs. CON -7.2±3.7%; P0.05). Neuromuscular electrical stimulation improves recovery from intensive training in professional team sports players. This strategy offers an easily applied recovery strategy which may have particular application during sleep and travel. Copyright © 2014 Sports Medicine Australia. Published by Elsevier Ltd. All rights reserved.

  11. Sodium butyrate stimulates cellular recovery from UV damage in xeroderma pigmentosum cells belonging to complementation group F

    International Nuclear Information System (INIS)

    Nishigori, Chikako; Takebe, Hiraku

    1987-01-01

    Possible stimulation of the DNA repair capacity by sodium butyrate in normal and xeroderma pigmentosum (XP) cells was investigated. XP cells belonging to the complementation group F showed considerable stimulation of DNA repair by sodium butyrate in terms of both the amount of unscheduled DNA synthesis (UDS) and the colony-forming ability after UV irradiation. UDS in XP cells belonging to the complementation group A was not enhanced, while normal cells showed slight enhancement, but less than that of XP F cells. In XP A, XP C, and normal cells, sodium butyrate treatment enhanced the killing effect of UV irradiation. The residual repair capacity in XP F cells appeared to be stimulated by sodium butyrate. (author)

  12. Requirement for the Phospho-H2AX Binding Module of Crb2 in Double-Strand Break Targeting and Checkpoint Activation▿

    Science.gov (United States)

    Sanders, Steven L.; Arida, Ahmad R.; Phan, Funita P.

    2010-01-01

    Activation of DNA damage checkpoints requires the rapid accumulation of numerous factors to sites of genomic lesions, and deciphering the mechanisms of this targeting is central to our understanding of DNA damage response. Histone modification has recently emerged as a critical element for the correct localization of damage response proteins, and one key player in this context is the fission yeast checkpoint mediator Crb2. Accumulation of Crb2 at ionizing irradiation-induced double-strand breaks (DSBs) requires two distinct histone marks, dimethylated H4 lysine 20 (H4K20me2) and phosphorylated H2AX (pH2AX). A tandem tudor motif in Crb2 directly binds H4K20me2, and this interaction is required for DSB targeting and checkpoint activation. Similarly, pH2AX is required for Crb2 localization to DSBs and checkpoint control. Crb2 can directly bind pH2AX through a pair of C-terminal BRCT repeats, but the functional significance of this binding has been unclear. Here we demonstrate that loss of its pH2AX-binding activity severely impairs the ability of Crb2 to accumulate at ionizing irradiation-induced DSBs, compromises checkpoint signaling, and disrupts checkpoint-mediated cell cycle arrest. These impairments are similar to that reported for abolition of pH2AX or mutation of the H4K20me2-binding tudor motif of Crb2. Intriguingly, a combined ablation of its two histone modification binding modules yields a strikingly additive reduction in Crb2 activity. These observations argue that binding of the Crb2 BRCT repeats to pH2AX is critical for checkpoint activity and provide new insight into the mechanisms of chromatin-mediated genome stability. PMID:20679488

  13. Requirement for the phospho-H2AX binding module of Crb2 in double-strand break targeting and checkpoint activation.

    Science.gov (United States)

    Sanders, Steven L; Arida, Ahmad R; Phan, Funita P

    2010-10-01

    Activation of DNA damage checkpoints requires the rapid accumulation of numerous factors to sites of genomic lesions, and deciphering the mechanisms of this targeting is central to our understanding of DNA damage response. Histone modification has recently emerged as a critical element for the correct localization of damage response proteins, and one key player in this context is the fission yeast checkpoint mediator Crb2. Accumulation of Crb2 at ionizing irradiation-induced double-strand breaks (DSBs) requires two distinct histone marks, dimethylated H4 lysine 20 (H4K20me2) and phosphorylated H2AX (pH2AX). A tandem tudor motif in Crb2 directly binds H4K20me2, and this interaction is required for DSB targeting and checkpoint activation. Similarly, pH2AX is required for Crb2 localization to DSBs and checkpoint control. Crb2 can directly bind pH2AX through a pair of C-terminal BRCT repeats, but the functional significance of this binding has been unclear. Here we demonstrate that loss of its pH2AX-binding activity severely impairs the ability of Crb2 to accumulate at ionizing irradiation-induced DSBs, compromises checkpoint signaling, and disrupts checkpoint-mediated cell cycle arrest. These impairments are similar to that reported for abolition of pH2AX or mutation of the H4K20me2-binding tudor motif of Crb2. Intriguingly, a combined ablation of its two histone modification binding modules yields a strikingly additive reduction in Crb2 activity. These observations argue that binding of the Crb2 BRCT repeats to pH2AX is critical for checkpoint activity and provide new insight into the mechanisms of chromatin-mediated genome stability.

  14. Anaphase onset before complete DNA replication with intact checkpoint responses

    DEFF Research Database (Denmark)

    Torres-Rosell, Jordi; De Piccoli, Giacomo; Cordon-Preciado, Violeta

    2007-01-01

    Cellular checkpoints prevent mitosis in the presence of stalled replication forks. Whether checkpoints also ensure the completion of DNA replication before mitosis is unknown. Here, we show that in yeast smc5-smc6 mutants, which are related to cohesin and condensin, replication is delayed, most...

  15. PD-1 Checkpoint Inhibitor Associated Autoimmune Encephalitis

    Directory of Open Access Journals (Sweden)

    Stephanie Schneider

    2017-05-01

    Full Text Available Objective: To report first-hand narrative experience of autoimmune encephalitis and to briefly review currently available evidence of autoimmune encephalitis in cancer patients treated with immune checkpoint inhibitors. Setting: A case study is presented on the management of a patient who developed autoimmune encephalitis during nivolumab monotherapy occurring after 28 weeks on anti-PD-1 monotherapy (nivolumab 3 mg/kg every 2 weeks for non-small cell lung cancer. Results: No substantial improvement was observed by antiepileptic treatment. After administration of 80 mg methylprednisolone, neurologic symptoms disappeared within 24 h and the patient fully recovered. Conclusions: Immune checkpoint inhibitor treatment can lead to autoimmune encephalitis. Clinical trial data indicate a frequency of autoimmune encephalitis of ≥0.1 to <1% with a higher probability during combined or sequential anti-CTLA-4/anti-PD-1 therapy than during anti-PD-1 or anti-PD-L1 monotherapy. Further collection of evidence and translational research is warranted.

  16. PD-1 checkpoint inhibition: Toxicities and management.

    Science.gov (United States)

    Hahn, Andrew W; Gill, David M; Agarwal, Neeraj; Maughan, Benjamin L

    2017-12-01

    With the recent approval of 5 PD-1/PD-L1 inhibitors for a number of malignancies, PD-1 axis inhibition is drastically changing the treatment landscape of immunotherapy in cancer. As PD-1/PD-L1 are involved in peripheral immune tolerance, inhibition of this immune checkpoint has led to novel immune-related adverse events including colitis, hepatitis, pneumonitis, rash, and endocrinopathies among many others. In this seminar, we will analyze the incidence of immune-related adverse events for nivolumab, pembrolizumab, atezolizumab, durvalumab, and avelumab. Then, we will discuss the specific management of the most common immune-mediated adverse events including colitis, hepatitis, pneumonitis, rash, endocrinopathies, nephritis, and neurologic toxicities. Immune-related adverse events are frequently treated with immunosuppressive medication such as steroids and mycofenolate mofetil. There are specific immune-related adverse events which are frequently seen by the treating oncologist from checkpoint inhibitors. It is essential to understand the recommended treatment options to minimize toxicity and mortality from this important class of anti-neoplastic therapies. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Laser-induced damage of fused silica optics at 355 nm due to backward stimulated Brillouin scattering: experimental and theoretical results.

    Science.gov (United States)

    Lamaignère, Laurent; Gaudfrin, Kévin; Donval, Thierry; Natoli, Jeanyves; Sajer, Jean-Michel; Penninckx, Denis; Courchinoux, Roger; Diaz, Romain

    2018-04-30

    Forward pump pulses with nanosecond duration are able to generate an acoustic wave via electrostriction through a few centimeters of bulk silica. Part of the incident energy is then scattered back on this sound wave, creating a backward Stokes pulse. This phenomenon known as stimulated Brillouin scattering (SBS) might induce first energy-loss, variable change of the temporal waveform depending on the location in the spatial profile making accurate metrology impossible, and moreover it might also initiate front surface damage making the optics unusable. Experiments performed on thick fused silica optics at 355 nm with single longitudinal mode pulses allowed us to detect, observe and quantify these backward pulses. Experimental results are first compared to theoretical calculations in order to strengthen our confidence in metrology. On this basis a phase-modulator has been implemented on the continuous-wave seeders of the lasers leading to pulses with a wide spectrum that suppress SBS and do not exhibit temporal overshoots that also reduce Kerr effects. The developed set-ups are used to check the reduction of the backward stimulated Brillouin scattering and they allow measuring with accuracy the rear surface damage of thick fused silica optics.

  18. Newly Emerging Immune Checkpoints: Promises for Future Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Robert J. Torphy

    2017-12-01

    Full Text Available Cancer immunotherapy has been a great breakthrough, with immune checkpoint inhibitors leading the way. Despite the clinical effectiveness of certain immune checkpoint inhibitors, the overall response rate remains low, and the effectiveness of immunotherapies for many tumors has been disappointing. There is substantial interest in looking for additional immune checkpoint molecules that may act as therapeutic targets for cancer. Recent advances during the last decade have identified several novel immune checkpoint targets, including lymphocyte activation gene-3 (LAG-3, B and T lymphocyte attenuator (BTLA, programmed death-1 homolog (PD-1H, T-cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIM-3/carcinoembryonic antigen cell adhesion molecule 1 (CEACAM1, and the poliovirus receptor (PVR-like receptors. The investigations into these molecules have generated promising results in preclinical studies. Herein, we will summarize our current progress and understanding of these newly-characterized immune checkpoints and their potential application in cancer immunotherapy.

  19. Gefarnate stimulates mucin-like glycoprotein secretion in conjunctival tissue and ameliorates corneal epithelial damage in animal dry-eye models

    Directory of Open Access Journals (Sweden)

    Dota A

    2013-01-01

    Full Text Available Atsuyoshi Dota, Yuko Takaoka-Shichijo, Masatsugu NakamuraOphthalmic Research and Development Center, Santen Pharmaceutical Co, Ltd, Ikoma-shi, Nara, JapanPurpose: The aim of this study was to evaluate the effect of gefarnate on mucin-like glycoprotein secretion in isolated rabbit conjunctival tissue, and on corneal epithelial damage in rabbit and cat dry-eye models.Methods: Conjunctival tissue isolated from rabbits was treated with gefarnate. Mucin-like glycoprotein was detected in the culture supernatant by an enzyme-linked lectin assay. Gefarnate ointment was topically applied to eyes once daily for 7 days in the rabbit dry-eye model, in which the lacrimal glands, Harderian gland, and nictitating membrane were removed, or for 4 weeks in the cat dry-eye model, in which the lacrimal gland and nictitating membrane were removed. Corneal epithelial damage was evaluated by measurement of corneal permeability by rose bengal in the rabbit model or by fluorescein staining in the cat model.Results: Gefarnate stimulated mucin-like glycoprotein secretion in conjunctival tissue in a dose-dependent manner. In the rabbit dry-eye model, application of gefarnate ointment to the eyes resulted in a dose-dependent decrease in rose bengal permeability in the cornea, with the effect being significant at concentrations of ≥0.3%. In the cat dry-eye model, application of gefarnate ointment resulted in a significant decrease in the corneal fluorescein staining score.Conclusion: These results suggest that gefarnate stimulates in vitro secretion of mucin-like glycoprotein in conjunctival tissue and ameliorates corneal epithelial damage in animal dry-eye models. Gefarnate may therefore be effective for treating dry eye.Keywords: gefarnate, fluorescein staining, rose bengal permeability, rabbit, cat, dry eye

  20. Immune checkpoint blockade therapy: The 2014 Tang prize in biopharmaceutical science

    Directory of Open Access Journals (Sweden)

    Ya-Shan Chen

    2015-02-01

    Full Text Available The first Tang Prize for Biopharmaceutical Science has been awarded to Prof. James P. Allison and Prof. Tasuku Honjo for their contributions leading to an entirely new way to treat cancer by blocking the molecules cytotoxic T lymphocyte-associated antigen 4 (CTLA-4 and programmed cell death protein 1 (PD-1 that turn off immune response. The treatment, called "immune checkpoint blockade therapy," has opened a new therapeutic era. Here the discoveries of the immune checkpoints and how they contribute to the maintenance of self-tolerance, as well as how to protect tissues from the excess immune responses causing damage are reviewed. The efforts made by Prof. Allison and Prof. Honjo for developing the most promising approaches to activate therapeutic antitumor immunity are also summarized. Since these certain immune checkpoint pathways appear to be one of the major mechanisms resulting in immune escape of tumors, the presence of anti-CTLA-4 and/or anti-PD-1 should contribute to removal of the inhibition signals for T cell activation. Subsequently, it will enhance specific T cell activation and, therefore, strengthen antitumor immunity.

  1. Cdk2 is required for p53-independent G2/M checkpoint control.

    Directory of Open Access Journals (Sweden)

    Jon H Chung

    2010-02-01

    Full Text Available The activation of phase-specific cyclin-dependent kinases (Cdks is associated with ordered cell cycle transitions. Among the mammalian Cdks, only Cdk1 is essential for somatic cell proliferation. Cdk1 can apparently substitute for Cdk2, Cdk4, and Cdk6, which are individually dispensable in mice. It is unclear if all functions of non-essential Cdks are fully redundant with Cdk1. Using a genetic approach, we show that Cdk2, the S-phase Cdk, uniquely controls the G(2/M checkpoint that prevents cells with damaged DNA from initiating mitosis. CDK2-nullizygous human cells exposed to ionizing radiation failed to exclude Cdk1 from the nucleus and exhibited a marked defect in G(2/M arrest that was unmasked by the disruption of P53. The DNA replication licensing protein Cdc6, which is normally stabilized by Cdk2, was physically associated with the checkpoint regulator ATR and was required for efficient ATR-Chk1-Cdc25A signaling. These findings demonstrate that Cdk2 maintains a balance of S-phase regulatory proteins and thereby coordinates subsequent p53-independent G(2/M checkpoint activation.

  2. Deficiency of vasodilator-stimulated phosphoprotein (VASP increases blood-brain-barrier damage and edema formation after ischemic stroke in mice.

    Directory of Open Access Journals (Sweden)

    Peter Kraft

    2010-12-01

    Full Text Available Stroke-induced brain edema formation is a frequent cause of secondary infarct growth and deterioration of neurological function. The molecular mechanisms underlying edema formation after stroke are largely unknown. Vasodilator-stimulated phosphoprotein (VASP is an important regulator of actin dynamics and stabilizes endothelial barriers through interaction with cell-cell contacts and focal adhesion sites. Hypoxia has been shown to foster vascular leakage by downregulation of VASP in vitro but the significance of VASP for regulating vascular permeability in the hypoxic brain in vivo awaits clarification.Focal cerebral ischemia was induced in Vasp(-/- mice and wild-type (WT littermates by transient middle cerebral artery occlusion (tMCAO. Evan's Blue tracer was applied to visualize the extent of blood-brain-barrier (BBB damage. Brain edema formation and infarct volumes were calculated from 2,3,5-triphenyltetrazolium chloride (TTC-stained brain slices. Both mouse groups were carefully controlled for anatomical and physiological parameters relevant for edema formation and stroke outcome. BBB damage (p0.05 towards worse neurological outcomes.Our study identifies VASP as critical regulator of BBB maintenance during acute ischemic stroke. Therapeutic modulation of VASP or VASP-dependent signalling pathways could become a novel strategy to combat excessive edema formation in ischemic brain damage.

  3. The tumor suppressor homolog in fission yeast, myh1+, displays a strong interaction with the checkpoint gene rad1+

    International Nuclear Information System (INIS)

    Jansson, Kristina; Warringer, Jonas; Farewell, Anne; Park, Han-Oh; Hoe, Kwang-Lae; Kim, Dong-Uk; Hayles, Jacqueline; Sunnerhagen, Per

    2008-01-01

    The DNA glycosylase MutY is strongly conserved in evolution, and homologs are found in most eukaryotes and prokaryotes examined. This protein is implicated in repair of oxidative DNA damage, in particular adenine mispaired opposite 7,8-dihydro-8-oxoguanine. Previous investigations in Escherichia coli, fission yeast, and mammalian cells show an association of mutations in MutY homologs with a mutator phenotype and carcinogenesis. Eukaryotic MutY homologs physically associate with several proteins with a role in replication, DNA repair, and checkpoint signaling, specifically the trimeric 9-1-1 complex. In a genetic investigation of the fission yeast MutY homolog, myh1 + , we show that the myh1 mutation confers a moderately increased UV sensitivity alone and in combination with mutations in several DNA repair genes. The myh1 rad1, and to a lesser degree myh1 rad9, double mutants display a synthetic interaction resulting in enhanced sensitivity to DNA damaging agents and hydroxyurea. UV irradiation of myh1 rad1 double mutants results in severe chromosome segregation defects and visible DNA fragmentation, and a failure to activate the checkpoint. Additionally, myh1 rad1 double mutants exhibit morphological defects in the absence of DNA damaging agents. We also found a moderate suppression of the slow growth and UV sensitivity of rhp51 mutants by the myh1 mutation. Our results implicate fission yeast Myh1 in repair of a wider range of DNA damage than previously thought, and functionally link it to the checkpoint pathway

  4. Neuromuscular complications of immune checkpoint inhibitor therapy.

    Science.gov (United States)

    Kolb, Noah A; Trevino, Christopher R; Waheed, Waqar; Sobhani, Fatemeh; Landry, Kara K; Thomas, Alissa A; Hehir, Mike

    2018-01-17

    Immune checkpoint inhibitor (ICPI) therapy unleashes the body's natural immune system to fight cancer. ICPIs improve overall cancer survival, however, the unbridling of the immune system may induce a variety of immune-related adverse events. Neuromuscular immune complications are rare but they can be severe. Myasthenia gravis and inflammatory neuropathy are the most common neuromuscular adverse events but a variety of others including inflammatory myopathy are reported. The pathophysiologic mechanism of these autoimmune disorders may differ from that of non-ICPI-related immune diseases. Accordingly, while the optimal treatment for ICPI-related neuromuscular disorders generally follows a traditional paradigm, there are important novel considerations in selecting appropriate immunosuppressive therapy. This review presents 2 new cases, a summary of neuromuscular ICPI complications, and an approach to the diagnosis and treatment of these disorders. Muscle Nerve, 2018. © 2018 Wiley Periodicals, Inc.

  5. Keeping checkpoint/restart viable for exascale systems.

    Energy Technology Data Exchange (ETDEWEB)

    Riesen, Rolf E.; Bridges, Patrick G. (IBM Research, Ireland, Mulhuddart, Dublin); Stearley, Jon R.; Laros, James H., III; Oldfield, Ron A.; Arnold, Dorian (University of New Mexico, Albuquerque, NM); Pedretti, Kevin Thomas Tauke; Ferreira, Kurt Brian; Brightwell, Ronald Brian

    2011-09-01

    Next-generation exascale systems, those capable of performing a quintillion (10{sup 18}) operations per second, are expected to be delivered in the next 8-10 years. These systems, which will be 1,000 times faster than current systems, will be of unprecedented scale. As these systems continue to grow in size, faults will become increasingly common, even over the course of small calculations. Therefore, issues such as fault tolerance and reliability will limit application scalability. Current techniques to ensure progress across faults like checkpoint/restart, the dominant fault tolerance mechanism for the last 25 years, are increasingly problematic at the scales of future systems due to their excessive overheads. In this work, we evaluate a number of techniques to decrease the overhead of checkpoint/restart and keep this method viable for future exascale systems. More specifically, this work evaluates state-machine replication to dramatically increase the checkpoint interval (the time between successive checkpoint) and hash-based, probabilistic incremental checkpointing using graphics processing units to decrease the checkpoint commit time (the time to save one checkpoint). Using a combination of empirical analysis, modeling, and simulation, we study the costs and benefits of these approaches on a wide range of parameters. These results, which cover of number of high-performance computing capability workloads, different failure distributions, hardware mean time to failures, and I/O bandwidths, show the potential benefits of these techniques for meeting the reliability demands of future exascale platforms.

  6. Prevention of DNA Rereplication Through a Meiotic Recombination Checkpoint Response

    Directory of Open Access Journals (Sweden)

    Nicole A. Najor

    2016-12-01

    Full Text Available In the budding yeast Saccharomyces cerevisiae, unnatural stabilization of the cyclin-dependent kinase inhibitor Sic1 during meiosis can trigger extra rounds of DNA replication. When programmed DNA double-strand breaks (DSBs are generated but not repaired due to absence of DMC1, a pathway involving the checkpoint gene RAD17 prevents this DNA rereplication. Further genetic analysis has now revealed that prevention of DNA rereplication also requires MEC1, which encodes a protein kinase that serves as a central checkpoint regulator in several pathways including the meiotic recombination checkpoint response. Downstream of MEC1, MEK1 is required through its function to inhibit repair between sister chromatids. By contrast, meiotic recombination checkpoint effectors that regulate gene expression and cyclin-dependent kinase activity are not necessary. Phosphorylation of histone H2A, which is catalyzed by Mec1 and the related Tel1 protein kinase in response to DSBs, and can help coordinate activation of the Rad53 checkpoint protein kinase in the mitotic cell cycle, is required for the full checkpoint response. Phosphorylation sites that are targeted by Rad53 in a mitotic S phase checkpoint response are also involved, based on the behavior of cells containing mutations in the DBF4 and SLD3 DNA replication genes. However, RAD53 does not appear to be required, nor does RAD9, which encodes a mediator of Rad53, consistent with their lack of function in the recombination checkpoint pathway that prevents meiotic progression. While this response is similar to a checkpoint mechanism that inhibits initiation of DNA replication in the mitotic cell cycle, the evidence points to a new variation on DNA replication control.

  7. Radiotherapy and immune checkpoint blockades: a snapshot in 2016

    Energy Technology Data Exchange (ETDEWEB)

    Koo, Tae Yool [Dept. of Radiation Oncology, Hallym University Chuncheon Sacred Heart Hospital, Chuncheon (Korea, Republic of); Kim, In Ah [Dept. of Radiation Oncology, Seoul National University College of Medicine, Seoul (Korea, Republic of)

    2016-12-15

    Immune checkpoint blockades including monoclonal antibodies (mAbs) of cytotoxic T-lymphocyte antigen-4 (CTLA-4), programmed death-1 (PD-1), and programmed death-ligand 1 (PD-L1) have been emerged as a promising anticancer therapy. Several immune checkpoint blockades have been approved by US Food and Drug Administration (FDA), and have shown notable success in clinical trials for patients with advanced melanoma and non-small cell lung cancer. Radiotherapy is a promising combination partner of immune checkpoint blockades due to its potent pro-immune effect. This review will cover the current issue and the future perspectives for combined with radiotherapy and immune checkpoint blockades based upon the available preclinical and clinical data.

  8. Correlated analysis of 2 MeV proton-induced radiation damage in CdZnTe crystals using photoluminescence and thermally stimulated current techniques

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Yaxu [State Key Laboratory of Solidification Processing, School of Materials Science and Engineering, Northwestern Polytechnical University, Xi’an 710072 (China); Key Laboratory of Radiation Detection Materials and Devices of Ministry of Industry and Information Technology, Northwestern Polytechnical University, Xi’an 710072 (China); Jie, Wanqi, E-mail: jwq@nwpu.edu.cn [State Key Laboratory of Solidification Processing, School of Materials Science and Engineering, Northwestern Polytechnical University, Xi’an 710072 (China); Key Laboratory of Radiation Detection Materials and Devices of Ministry of Industry and Information Technology, Northwestern Polytechnical University, Xi’an 710072 (China); Rong, Caicai [Institute of Modern Physics, Applied Ion Beam Physics Laboratory, Fudan University, Shanghai 200433 (China); Wang, Yuhan; Xu, Lingyan; Xu, Yadong [State Key Laboratory of Solidification Processing, School of Materials Science and Engineering, Northwestern Polytechnical University, Xi’an 710072 (China); Key Laboratory of Radiation Detection Materials and Devices of Ministry of Industry and Information Technology, Northwestern Polytechnical University, Xi’an 710072 (China); Lv, Haoyan; Shen, Hao [Institute of Modern Physics, Applied Ion Beam Physics Laboratory, Fudan University, Shanghai 200433 (China); Du, Guanghua [Materials Research Center, Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Fu, Xu [State Key Laboratory of Solidification Processing, School of Materials Science and Engineering, Northwestern Polytechnical University, Xi’an 710072 (China); Key Laboratory of Radiation Detection Materials and Devices of Ministry of Industry and Information Technology, Northwestern Polytechnical University, Xi’an 710072 (China); and others

    2016-11-01

    Highlights: • 2 MeV proton-induced radiation damage in CdZnTe crystals is investigated by PL and TSC techniques. • The influence of radiation damage on the luminescent and electrical properties of CdZnTe crystals is studied. • Intensity of PL spectrum is found to decrease significantly in irradiated regions, suggesting the increase of non-radiative recombination centers. • A correlated analysis of PL and TSC spectra suggests that the density of dislocations and A-centers increase after proton irradiation. - Abstract: Radiation damage induced by 2 MeV protons in CdZnTe crystals has been studied by means of photoluminescence (PL) and thermally stimulated current (TSC) techniques. A notable quenching of PL intensity is observed in the regions irradiated with a fluence of 6 × 10{sup 13} p/cm{sup 2}, suggesting the increase of non-radiative recombination centers. Moreover, the intensity of emission peak D{sub complex} centered at 1.48 eV dominates in the PL spectrum obtained from irradiated regions, ascribed to the increase of interstitial dislocation loops and A centers. The intensity of TSC spectra in irradiated regions decreases compared to the virgin regions, resulting from the charge collection inefficiency caused by proton-induced recombination centers. By comparing the intensity of identified traps obtained from numerical fitting using simultaneous multiple peak analysis (SIMPA) method, it suggests that proton irradiation under such dose can introduce high density of dislocation and A-centers in CdZnTe crystals, consistent with PL results.

  9. A New Adaptive Checkpointing Strategy for Mobile Computing

    Institute of Scientific and Technical Information of China (English)

    MENChaoguang; ZUODecheng; YANGXiaozong

    2005-01-01

    Adaptive checkpointing strategy is an efficient recovery scheme, which is suitable for mobile computing system. However, all existing adaptive checkpointing schemes are not correct to recover system when failure occurs in some special period. In this paper, the issues that will lead to system inconsistency are first discussed and then a new adaptive strategy that can recover system to correct consistent state is proposed. Our algorithm improves system recovery performance because only failure process needs rollback through logging.

  10. Alpha-Melanocyte Stimulating Hormone Protects against Cytokine-Induced Barrier Damage in Caco-2 Intestinal Epithelial Monolayers.

    Directory of Open Access Journals (Sweden)

    Judit Váradi

    Full Text Available Alpha-melanocyte-stimulating hormone (α-MSH is a potent anti-inflammatory peptide with cytoprotective effect in various tissues. The present investigation demonstrates the ability of α-MSH to interact with intestinal epithelial cell monolayers and mitigate inflammatory processes of the epithelial barrier. The protective effect of α-MSH was studied on Caco-2 human intestinal epithelial monolayers, which were disrupted by exposure to tumor necrosis factor-α and interleukin-1β. The barrier integrity was assessed by measuring transepithelial electric resistance (TEER and permeability for marker molecules. Caco-2 monolayers were evaluated by immunohistochemistry for expression of melanocortin-1 receptor and tight junction proteins ZO-1 and claudin-4. The activation of nuclear factor kappa beta (NF-κB was detected by fluorescence microscopy and inflammatory cytokine expression was assessed by flow cytometric bead array cytokine assay. Exposure of Caco-2 monolayers to proinflammatory cytokines lowered TEER and increased permeability for fluorescein and albumin, which was accompanied by changes in ZO-1 and claudin-4 immunostaining. α-MSH was able to prevent inflammation-associated decrease of TEER in a dose-dependent manner and reduce the increased permeability for paracellular marker fluorescein. Further immunohistochemistry analysis revealed proinflammatory cytokine induced translocation of the NF-κB p65 subunit into Caco-2 cell nuclei, which was inhibited by α-MSH. As a result the IL-6 and IL-8 production of Caco-2 monolayers were also decreased with different patterns by the addition of α-MSH to the culture medium. In conclusion, Caco-2 cells showed a positive immunostaining for melanocortin-1 receptor and α-MSH protected Caco-2 cells against inflammatory barrier dysfunction and inflammatory activation induced by tumor necrosis factor-α and interleukin-1β cytokines.

  11. Checkpoint blockade in combination with cancer vaccines.

    Science.gov (United States)

    Morse, Michael A; Lyerly, H Kim

    2015-12-16

    Checkpoint blockade, prevention of inhibitory signaling that limits activation or function of tumor antigen-specific T cells responses, is revolutionizing the treatment of many poor prognosis malignancies. Indeed monoclonal antibodies that modulate signaling through the inhibitory molecules CTLA-4 and PD-1 are now clinically available; however, many tumors, demonstrate minimal response suggesting the need for combinations with other therapeutic strategies. Because an inadequate frequency of activated tumor antigen-specific T cells in the tumor environment, the so-called non-inflamed phenotype, is observed in some malignancies, other rationale partners are modalities that lead to enhanced T cell activation (vaccines, cytokines, toll-like receptor agonists, and other anticancer therapies such as chemo-, radio- or targeted therapies that lead to release of antigen from tumors). This review will focus on preclinical and clinical data supporting the use of cancer vaccines with anti-CTLA-4 and anti-PD-1/PD-L1 antibodies. Preliminary preclinical data demonstrate enhanced antitumor activity although the results in human studies are less clear. Broader combinations of multiple immune modulators are now under study. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. Aurora-B Mediated ATM Serine 1403 Phosphorylation Is Required For Mitotic ATM Activation and the Spindle Checkpoint

    OpenAIRE

    Yang, Chunying; Tang, Xi; Guo, Xiaojing; Niikura, Yohei; Kitagawa, Katsumi; Cui, Kemi; Wong, Stephen T.C.; Fu, Li; Xu, Bo

    2011-01-01

    The ATM kinase plays a critical role in the maintenance of genetic stability. ATM is activated in response to DNA damage and is essential for cell cycle checkpoints. Here, we report that ATM is activated in mitosis in the absence of DNA damage. We demonstrate that mitotic ATM activation is dependent on the Aurora-B kinase and that Aurora-B phosphorylates ATM on serine 1403. This phosphorylation event is required for mitotic ATM activation. Further, we show that loss of ATM function results in...

  13. The Checkpoint Kinase 1 Inhibitor Prexasertib Induces Regression of Preclinical Models of Human Neuroblastoma.

    Science.gov (United States)

    Lowery, Caitlin D; VanWye, Alle B; Dowless, Michele; Blosser, Wayne; Falcon, Beverly L; Stewart, Julie; Stephens, Jennifer; Beckmann, Richard P; Bence Lin, Aimee; Stancato, Louis F

    2017-08-01

    Purpose: Checkpoint kinase 1 (CHK1) is a key regulator of the DNA damage response and a mediator of replication stress through modulation of replication fork licensing and activation of S and G 2 -M cell-cycle checkpoints. We evaluated prexasertib (LY2606368), a small-molecule CHK1 inhibitor currently in clinical testing, in multiple preclinical models of pediatric cancer. Following an initial assessment of prexasertib activity, this study focused on the preclinical models of neuroblastoma. Experimental Design: We evaluated the antiproliferative activity of prexasertib in a panel of cancer cell lines; neuroblastoma cell lines were among the most sensitive. Subsequent Western blot and immunofluorescence analyses measured DNA damage and DNA repair protein activation. Prexasertib was investigated in several cell line-derived xenograft mouse models of neuroblastoma. Results: Within 24 hours, single-agent prexasertib promoted γH2AX-positive double-strand DNA breaks and phosphorylation of DNA damage sensors ATM and DNA-PKcs, leading to neuroblastoma cell death. Knockdown of CHK1 and/or CHK2 by siRNA verified that the double-strand DNA breaks and cell death elicited by prexasertib were due to specific CHK1 inhibition. Neuroblastoma xenografts rapidly regressed following prexasertib administration, independent of starting tumor volume. Decreased Ki67 and increased immunostaining of endothelial and pericyte markers were observed in xenografts after only 6 days of exposure to prexasertib, potentially indicating a swift reduction in tumor volume and/or a direct effect on tumor vasculature. Conclusions: Overall, these data demonstrate that prexasertib is a specific inhibitor of CHK1 in neuroblastoma and leads to DNA damage and cell death in preclinical models of this devastating pediatric malignancy. Clin Cancer Res; 23(15); 4354-63. ©2017 AACR . ©2017 American Association for Cancer Research.

  14. Replicative stress and alterations in cell cycle checkpoint controls following acetaminophen hepatotoxicity restrict liver regeneration.

    Science.gov (United States)

    Viswanathan, Preeti; Sharma, Yogeshwar; Gupta, Priya; Gupta, Sanjeev

    2018-03-05

    Acetaminophen hepatotoxicity is a leading cause of hepatic failure with impairments in liver regeneration producing significant mortality. Multiple intracellular events, including oxidative stress, mitochondrial damage, inflammation, etc., signify acetaminophen toxicity, although how these may alter cell cycle controls has been unknown and was studied for its significance in liver regeneration. Assays were performed in HuH-7 human hepatocellular carcinoma cells, primary human hepatocytes and tissue samples from people with acetaminophen-induced acute liver failure. Cellular oxidative stress, DNA damage and cell proliferation events were investigated by mitochondrial membrane potential assays, flow cytometry, fluorescence staining, comet assays and spotted arrays for protein expression after acetaminophen exposures. In experimental groups with acetaminophen toxicity, impaired mitochondrial viability and substantial DNA damage were observed with rapid loss of cells in S and G2/M and cell cycle restrictions or even exit in the remainder. This resulted from altered expression of the DNA damage regulator, ATM and downstream transducers, which imposed G1/S checkpoint arrest, delayed entry into S and restricted G2 transit. Tissues from people with acute liver failure confirmed hepatic DNA damage and cell cycle-related lesions, including restrictions of hepatocytes in aneuploid states. Remarkably, treatment of cells with a cytoprotective cytokine reversed acetaminophen-induced restrictions to restore cycling. Cell cycle lesions following mitochondrial and DNA damage led to failure of hepatic regeneration in acetaminophen toxicity but their reversibility offers molecular targets for treating acute liver failure. © 2018 John Wiley & Sons Ltd.

  15. Berkeley lab checkpoint/restart (BLCR) for Linux clusters

    International Nuclear Information System (INIS)

    Hargrove, Paul H; Duell, Jason C

    2006-01-01

    This article describes the motivation, design and implementation of Berkeley Lab Checkpoint/Restart (BLCR), a system-level checkpoint/restart implementation for Linux clusters that targets the space of typical High Performance Computing applications, including MPI. Application-level solutions, including both checkpointing and fault-tolerant algorithms, are recognized as more time and space efficient than system-level checkpoints, which cannot make use of any application-specific knowledge. However, system-level checkpointing allows for preemption, making it suitable for responding to ''fault precursors'' (for instance, elevated error rates from ECC memory or network CRCs, or elevated temperature from sensors). Preemption can also increase the efficiency of batch scheduling; for instance reducing idle cycles (by allowing for shutdown without any queue draining period or reallocation of resources to eliminate idle nodes when better fitting jobs are queued), and reducing the average queued time (by limiting large jobs to running during off-peak hours, without the need to limit the length of such jobs). Each of these potential uses makes BLCR a valuable tool for efficient resource management in Linux clusters

  16. Enhanced granulocyte growth on peritoneal cell-coated membranes following irradiation: a dual effect of humoral stimulation and repair of x ray-induced damage to the microenvironment

    International Nuclear Information System (INIS)

    Turner, A.R.; Pfrimmer, W.J.; Boggs, D.R.; Carpe, A.I.

    1978-01-01

    An experimental model of the hematopoietic microenvironment was created by allowing a peritoneal cell coating to form on a disk of cellulose acetate placed in the peritoneal cavity of mice. An effective microenvironment capable of supporting colony growth, primarily granulocytic, was established if the cellulose acetate disk was in the peritoneum for 3 to 5 days. Its effectiveness was hampered by transferring it to another mouse or by exposure to toxic agents such as a propylene glycol-ethanol mixture or irradiation. An exponential dose-related decrease in colony formation was seen with increasing doses or irradiation of the microenvironment before colonization. After a low dose of irradiation, recovery of colony support capacity occurred over a 6-day period. Enhancement of colony growth was seen when cell injection was delayed for 2 to 3 days after irradiation. The effects of irradiation on the cellular stroma were separated from the systemic changes in the host by transferring an established hematopoietic microenvironment to a secondary host. It was shown that there are two distinct effects of irradiation on granulocytic colony growth; one was a short-lived period, 2 to 3 days of stimulation, presumably humoral, and the other was dose-dependent reversible microenvironment damage

  17. Developmental checkpoints and feedback circuits time insect maturation

    DEFF Research Database (Denmark)

    Rewitz, Kim Furbo; Yamanaka, Naoki; O'Connor, Michael B.

    2013-01-01

    as external cues, to time production and release of ecdysone. Based on results discussed here, we suggest that developmental progression to adulthood is controlled by checkpoints that regulate the genetic timing program enabling it to adapt to different environmental conditions. These checkpoints utilize...... a number of signaling pathways to modulate ecdysone production in the prothoracic gland. Release of ecdysone activates an autonomous cascade of both feedforward and feedback signals that determine the duration of the ecdysone pulse at each developmental transitions. Conservation of the genetic mechanisms...... that coordinate the juvenile-adult transition suggests that insights from the fruit fly Drosophila will provide a framework for future investigation of developmental timing in metazoans....

  18. The Dynamical Mechanisms of the Cell Cycle Size Checkpoint

    International Nuclear Information System (INIS)

    Feng Shi-Fu; Yang Ling; Yan Jie; Liu Zeng-Rong

    2012-01-01

    Cell division must be tightly coupled to cell growth in order to maintain cell size, whereas the mechanisms of how initialization of mitosis is regulated by cell size remain to be elucidated. We develop a mathematical model of the cell cycle, which incorporates cell growth to investigate the dynamical properties of the size checkpoint in embryos of Xenopus laevis. We show that the size checkpoint is naturally raised from a saddle-node bifurcation, and in a mutant case, the cell loses its size control ability due to the loss of this saddle-node point

  19. Rad9 contribution to radiosensitivity and the G2 checkpoint in a DT40 cell line

    Energy Technology Data Exchange (ETDEWEB)

    Kumano, Tomoyasu [Kanazawa Univ. (Japan). Graduate School of Medical Science

    2002-12-01

    In fission yeast, the rad9 (radiation sensitive) gene was cloned from a mutant that is sensitive to ionizing radiation, ultraviolet and hydroxyurea. This gene has also been shown to be required for a DNA damage checkpoint. Orthologues of the rad9 gene have recently been identified in higher eukaryote cells including human. Here we generated Rad9 knockout (Rad9-/-) cells from the chicken B lymphocyte line DT40 to examine the role of Rad9 in higher eukaryotes. First we isolated a part of the chicken Rad9 gene which was 54% identical with human Rad9 at the amino acid sequence level. Next we isolated genomic clones, determined exons and introns, and constructed targeting vectors designed to disrupt exon 1-3 of the chicken Rad9 gene by replacement with a drug-resistant gene. Successful targeted integration was verified by Southern blot analysis and the disruption of the Rad9 gene was confirmed by reverse transcription polymerase chain reaction (RT-PCR). To analyze the radiosensitivity of these Rad9-/- cells, we monitored the clonogenic survival after various degrees of X-ray irradiation. Rad9-/- cells were more sensitive to X-rays than wild type cells at all dosages. However, these cells were less sensitive than ATM knockout (ATM-/-) cells that are known to be X-ray sensitive and that showed a defective checkpoint control. In contrast, Rad9-/- cells were markedly more sensitive to ultraviolet and hydroxyruea. In addition, we assessed the G2 checkpoint by measurement of the mitotic index that is the fraction of the accumulating number of cells in mitosis at various times after X-ray irradiation. While the number of mitotic wild type cells did not increase until 2 hrs after X-ray irradiation, the number of mitotic Rad9-/- cells showed an increase similar to that of ATM-/- cells. These results suggest that just as in fission yeast, in higher eukaryotes Rad9 also contributes to X-ray, ultraviolet and hydroxyurea sensitivity, and plays an important role in the G2 checkpoint

  20. Rad9 contribution to radiosensitivity and the G2 checkpoint in a DT40 cell line

    International Nuclear Information System (INIS)

    Kumano, Tomoyasu

    2002-01-01

    In fission yeast, the rad9 (radiation sensitive) gene was cloned from a mutant that is sensitive to ionizing radiation, ultraviolet and hydroxyurea. This gene has also been shown to be required for a DNA damage checkpoint. Orthologues of the rad9 gene have recently been identified in higher eukaryote cells including human. Here we generated Rad9 knockout (Rad9-/-) cells from the chicken B lymphocyte line DT40 to examine the role of Rad9 in higher eukaryotes. First we isolated a part of the chicken Rad9 gene which was 54% identical with human Rad9 at the amino acid sequence level. Next we isolated genomic clones, determined exons and introns, and constructed targeting vectors designed to disrupt exon 1-3 of the chicken Rad9 gene by replacement with a drug-resistant gene. Successful targeted integration was verified by Southern blot analysis and the disruption of the Rad9 gene was confirmed by reverse transcription polymerase chain reaction (RT-PCR). To analyze the radiosensitivity of these Rad9-/- cells, we monitored the clonogenic survival after various degrees of X-ray irradiation. Rad9-/- cells were more sensitive to X-rays than wild type cells at all dosages. However, these cells were less sensitive than ATM knockout (ATM-/-) cells that are known to be X-ray sensitive and that showed a defective checkpoint control. In contrast, Rad9-/- cells were markedly more sensitive to ultraviolet and hydroxyruea. In addition, we assessed the G2 checkpoint by measurement of the mitotic index that is the fraction of the accumulating number of cells in mitosis at various times after X-ray irradiation. While the number of mitotic wild type cells did not increase until 2 hrs after X-ray irradiation, the number of mitotic Rad9-/- cells showed an increase similar to that of ATM-/- cells. These results suggest that just as in fission yeast, in higher eukaryotes Rad9 also contributes to X-ray, ultraviolet and hydroxyurea sensitivity, and plays an important role in the G2 checkpoint

  1. The DNA Replication Checkpoint Directly Regulates MBF-Dependent G1/S Transcription▿

    OpenAIRE

    Dutta, Chaitali; Patel, Prasanta K.; Rosebrock, Adam; Oliva, Anna; Leatherwood, Janet; Rhind, Nicholas

    2008-01-01

    The DNA replication checkpoint transcriptionally upregulates genes that allow cells to adapt to and survive replication stress. Our results show that, in the fission yeast Schizosaccharomyces pombe, the replication checkpoint regulates the entire G1/S transcriptional program by directly regulating MBF, the G1/S transcription factor. Instead of initiating a checkpoint-specific transcriptional program, the replication checkpoint targets MBF to maintain the normal G1/S transcriptional program du...

  2. Development of cell-cycle checkpoint therapy for solid tumors.

    Science.gov (United States)

    Tamura, Kenji

    2015-12-01

    Cellular proliferation is tightly controlled by several cell-cycle checkpoint proteins. In cancer, the genes encoding these proteins are often disrupted and cause unrestrained cancer growth. The proteins are over-expressed in many malignancies; thus, they are potential targets for anti-cancer therapies. These proteins include cyclin-dependent kinase, checkpoint kinase, WEE1 kinase, aurora kinase and polo-like kinase. Cyclin-dependent kinase inhibitors are the most advanced cell-cycle checkpoint therapeutics available. For instance, palbociclib (PD0332991) is a first-in-class, oral, highly selective inhibitor of CDK4/6 and, in combination with letrozole (Phase II; PALOMA-1) or with fulvestrant (Phase III; PALOMA-3), it has significantly prolonged progression-free survival, in patients with metastatic estrogen receptor-positive, HER2-negative breast cancer, in comparison with that observed in patients using letrozole, or fulvestrant alone, respectively. In this review, we provide an overview of the current compounds available for cell-cycle checkpoint protein-directed therapy for solid tumors. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  3. Immune-Checkpoint Blockade and Active Immunotherapy for Glioma

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, Brian J. [Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Brain Tumor Program, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213 (United States); Pollack, Ian F. [Brain Tumor Program, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213 (United States); Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Okada, Hideho, E-mail: okadah@upmc.edu [Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Brain Tumor Program, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213 (United States); Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States); Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213 (United States)

    2013-11-01

    Cancer immunotherapy has made tremendous progress, including promising results in patients with malignant gliomas. Nonetheless, the immunological microenvironment of the brain and tumors arising therein is still believed to be suboptimal for sufficient antitumor immune responses for a variety of reasons, including the operation of “immune-checkpoint” mechanisms. While these mechanisms prevent autoimmunity in physiological conditions, malignant tumors, including brain tumors, actively employ these mechanisms to evade from immunological attacks. Development of agents designed to unblock these checkpoint steps is currently one of the most active areas of cancer research. In this review, we summarize recent progresses in the field of brain tumor immunology with particular foci in the area of immune-checkpoint mechanisms and development of active immunotherapy strategies. In the last decade, a number of specific monoclonal antibodies designed to block immune-checkpoint mechanisms have been developed and show efficacy in other cancers, such as melanoma. On the other hand, active immunotherapy approaches, such as vaccines, have shown encouraging outcomes. We believe that development of effective immunotherapy approaches should ultimately integrate those checkpoint-blockade agents to enhance the efficacy of therapeutic approaches. With these agents available, it is going to be quite an exciting time in the field. The eventual success of immunotherapies for brain tumors will be dependent upon not only an in-depth understanding of immunology behind the brain and brain tumors, but also collaboration and teamwork for the development of novel trials that address multiple layers of immunological challenges in gliomas.

  4. Immune checkpoint inhibitors for nonsmall cell lung cancer treatment

    Directory of Open Access Journals (Sweden)

    Yuh-Min Chen

    2017-01-01

    Full Text Available Immune checkpoint inhibition with blocking antibodies that target cytotoxic T-lymphocyte antigen-4 (CTLA-4 and the programmed cell death protein 1 (PD-1 pathway [PD-1/programmed death-ligand 1 (PD-L1] have demonstrated promise in a variety of malignancies. While ipilimumab has been approved as a CTLA-4 blocking antibody by the US Food and Drug Administration for the treatment of advanced melanoma, it is still not approved for lung cancer treatment. In contrast, nivolumab and pembrolizumab, both PD-1 blocking antibodies, have been approved for second-line treatment of nonsmall cell lung cancer in 2015 because of their high potency and long-lasting effects in some patient subgroups. Other PD-1 and PD-L1 monoclonal antibodies are also in active development phase. Treatment with such immune checkpoint inhibitors is associated with a unique pattern of immune-related adverse events or side effects. Combination approaches involving CTLA-4 and PD-1/PD-L1 blockade or checkpoint inhibitors with chemotherapy or radiotherapy are being investigated to determine whether they may enhance the efficacy of treatment. Despite many challenges ahead, immunotherapy with checkpoint inhibitors has already become a new and important treatment modality for lung cancer in the last decade following the discovery of targeted therapy.

  5. Immune-Checkpoint Blockade and Active Immunotherapy for Glioma

    International Nuclear Information System (INIS)

    Ahn, Brian J.; Pollack, Ian F.; Okada, Hideho

    2013-01-01

    Cancer immunotherapy has made tremendous progress, including promising results in patients with malignant gliomas. Nonetheless, the immunological microenvironment of the brain and tumors arising therein is still believed to be suboptimal for sufficient antitumor immune responses for a variety of reasons, including the operation of “immune-checkpoint” mechanisms. While these mechanisms prevent autoimmunity in physiological conditions, malignant tumors, including brain tumors, actively employ these mechanisms to evade from immunological attacks. Development of agents designed to unblock these checkpoint steps is currently one of the most active areas of cancer research. In this review, we summarize recent progresses in the field of brain tumor immunology with particular foci in the area of immune-checkpoint mechanisms and development of active immunotherapy strategies. In the last decade, a number of specific monoclonal antibodies designed to block immune-checkpoint mechanisms have been developed and show efficacy in other cancers, such as melanoma. On the other hand, active immunotherapy approaches, such as vaccines, have shown encouraging outcomes. We believe that development of effective immunotherapy approaches should ultimately integrate those checkpoint-blockade agents to enhance the efficacy of therapeutic approaches. With these agents available, it is going to be quite an exciting time in the field. The eventual success of immunotherapies for brain tumors will be dependent upon not only an in-depth understanding of immunology behind the brain and brain tumors, but also collaboration and teamwork for the development of novel trials that address multiple layers of immunological challenges in gliomas

  6. A checkpoint compression study for high-performance computing systems

    Energy Technology Data Exchange (ETDEWEB)

    Ibtesham, Dewan [Univ. of New Mexico, Albuquerque, NM (United States). Dept. of Computer Science; Ferreira, Kurt B. [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States). Scalable System Software Dept.; Arnold, Dorian [Univ. of New Mexico, Albuquerque, NM (United States). Dept. of Computer Science

    2015-02-17

    As high-performance computing systems continue to increase in size and complexity, higher failure rates and increased overheads for checkpoint/restart (CR) protocols have raised concerns about the practical viability of CR protocols for future systems. Previously, compression has proven to be a viable approach for reducing checkpoint data volumes and, thereby, reducing CR protocol overhead leading to improved application performance. In this article, we further explore compression-based CR optimization by exploring its baseline performance and scaling properties, evaluating whether improved compression algorithms might lead to even better application performance and comparing checkpoint compression against and alongside other software- and hardware-based optimizations. Our results highlights are: (1) compression is a very viable CR optimization; (2) generic, text-based compression algorithms appear to perform near optimally for checkpoint data compression and faster compression algorithms will not lead to better application performance; (3) compression-based optimizations fare well against and alongside other software-based optimizations; and (4) while hardware-based optimizations outperform software-based ones, they are not as cost effective.

  7. CHECKPOINT INHIBITOR IMMUNE THERAPY: Systemic Indications and Ophthalmic Side Effects.

    Science.gov (United States)

    Dalvin, Lauren A; Shields, Carol L; Orloff, Marlana; Sato, Takami; Shields, Jerry A

    2018-06-01

    To review immune checkpoint inhibitor indications and ophthalmic side effects. A literature review was performed using a PubMed search for publications between 1990 and 2017. Immune checkpoint inhibitors are designed to treat system malignancies by targeting one of three ligands, leading to T-cell activation for attack against malignant cells. These ligands (and targeted drug) include cytotoxic T-lymphocyte antigen-4 (CTLA-4, ipilimumab), programmed death protein 1 (PD-1, pembrolizumab, nivolumab), and programmed death ligand-1 (PD-L1, atezolizumab, avelumab, durvalumab). These medications upregulate the immune system and cause autoimmune-like side effects. Ophthalmic side effects most frequently manifest as uveitis (1%) and dry eye (1-24%). Other side effects include myasthenia gravis (n = 19 reports), inflammatory orbitopathy (n = 11), keratitis (n = 3), cranial nerve palsy (n = 3), optic neuropathy (n = 2), serous retinal detachment (n = 2), extraocular muscle myopathy (n = 1), atypical chorioretinal lesions (n = 1), immune retinopathy (n = 1), and neuroretinitis (n = 1). Most inflammatory side effects are managed with topical or periocular corticosteroids, but advanced cases require systemic corticosteroids and cessation of checkpoint inhibitor therapy. Checkpoint inhibitors enhance the immune system by releasing inhibition on T cells, with risk of autoimmune-like side effects. Ophthalmologists should include immune-related adverse events in their differential when examining cancer patients with new ocular symptoms.

  8. Checkpointing for graceful degradation in distributed embedded systems

    Science.gov (United States)

    Sababha, Belal Hussein

    Graceful degradation is an approach to developing dependable safety-critical embedded applications, where redundant active or standby resources are used to cope with faults through a system reconfiguration at run-time. Compared to traditional hardware and software redundancy, it is a promising technique that may achieve dependability with a significant reduction in cost, size, weight, and power requirements. Reconfiguration at run-time necessitates using proper checkpointing protocols to support state reservation to ensure correct task restarts after a system reconfiguration. One of the most common checkpointing protocols are communication induced checkpointing (CIC) protocols, which are well developed and understood for large parallel and information systems, but not much has been done for resource limited embedded systems. This work implements and evaluates some of the most common CIC protocols in a periodic resource constrained distributed embedded system for graceful degradation purposes. A test-bed has been developed and used for the evaluation of the various protocols. The implemented protocols are thoroughly studied and performances are contrasted. Specifically the periodicity property and how it benefits checkpointing in embedded systems is investigated. This work introduces a unique effort of CIC protocol implementation and evaluation in the field of distributed embedded systems. Other than providing a test-bed for graceful degradation support, this work shows that some checkpointing protocols that are not efficient in large information systems and supercomputers perform well in embedded systems. We show that a simple index-based CIC protocol, such as the BCS protocol, is more appropriate in embedded system applications compared to other protocols that piggyback a significant amount of information to reduce the number of forced checkpoints. Finally, this work proposes a whole graceful degradation approach to achieve fault tolerance in resource constrained

  9. Checkpoint inhibitors in endometrial cancer: preclinical rationale and clinical activity.

    Science.gov (United States)

    Mittica, Gloria; Ghisoni, Eleonora; Giannone, Gaia; Aglietta, Massimo; Genta, Sofia; Valabrega, Giorgio

    2017-10-27

    Treatment of advanced and recurrent endometrial cancer (EC) is still an unmet need for oncologists and gynecologic oncologists. The Cancer Genome Atlas Research Network (TCGA) recently provided a new genomic classification, dividing EC in four subgroups. Two types of EC, the polymerase epsilon (POLE)-ultra-mutated and the microsatellite instability-hyper-mutated (MSI-H), are characterized by a high mutation rate providing the rationale for a potential activity of checkpoint inhibitors. We analyzed all available evidence supporting the role of tumor microenvironment (TME) in EC development and the therapeutic implications offered by immune checkpoint inhibitors in this setting. We performed a review on Pubmed with Mesh keywords 'endometrial cancer' and the name of each checkpoint inhibitor discussed in the article. The same search was operated on clinicaltrial.gov to identify ongoing clinical trials exploring PD-1/PD-L1 and CTLA-4 axis in EC, particularly focusing on POLE-ultra-muted and MSI-H cancer types. POLE-ultra-mutated and MSI-H ECs showed an active TME expressing high number of neo-antigens and an elevated amount of tumor infiltrating lymphocytes (TILs). Preliminary results from a phase-1 clinical trial (KEYNOTE-028) demonstrated antitumor activity of Pembrolizumab in EC. Moreover, both Pembrolizumab and Nivolumab reported durable clinical responses in POLE-ultra-mutated patients. Immune checkpoint inhibitors are an attractive option in POLE-ultra-mutated and MSI-H ECs. Future investigations in these subgroups include combinations of checkpoints inhibitors with chemotherapy and small tyrosine kinase inhibitors (TKIs) to enhance a more robust intra-tumoral immune response.

  10. Cell size checkpoint control by the retinoblastoma tumor suppressor pathway.

    Science.gov (United States)

    Fang, Su-Chiung; de los Reyes, Chris; Umen, James G

    2006-10-13

    Size control is essential for all proliferating cells, and is thought to be regulated by checkpoints that couple cell size to cell cycle progression. The aberrant cell-size phenotypes caused by mutations in the retinoblastoma (RB) tumor suppressor pathway are consistent with a role in size checkpoint control, but indirect effects on size caused by altered cell cycle kinetics are difficult to rule out. The multiple fission cell cycle of the unicellular alga Chlamydomonas reinhardtii uncouples growth from division, allowing direct assessment of the relationship between size phenotypes and checkpoint function. Mutations in the C. reinhardtii RB homolog encoded by MAT3 cause supernumerous cell divisions and small cells, suggesting a role for MAT3 in size control. We identified suppressors of an mat3 null allele that had recessive mutations in DP1 or dominant mutations in E2F1, loci encoding homologs of a heterodimeric transcription factor that is targeted by RB-related proteins. Significantly, we determined that the dp1 and e2f1 phenotypes were caused by defects in size checkpoint control and were not due to a lengthened cell cycle. Despite their cell division defects, mat3, dp1, and e2f1 mutants showed almost no changes in periodic transcription of genes induced during S phase and mitosis, many of which are conserved targets of the RB pathway. Conversely, we found that regulation of cell size was unaffected when S phase and mitotic transcription were inhibited. Our data provide direct evidence that the RB pathway mediates cell size checkpoint control and suggest that such control is not directly coupled to the magnitude of periodic cell cycle transcription.

  11. Fission yeast strains with circular chromosomes require the 9-1-1 checkpoint complex for the viability in response to the anti-cancer drug 5-fluorodeoxyuridine.

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    Hossain Mohammad Shamim

    Full Text Available Thymidine kinase converts 5-fluorodeoxyuridine to 5-fluorodeoxyuridine monophosphate, which causes disruption of deoxynucleotide triphosphate ratios. The fission yeast Schizosaccharomyces pombe does not express endogenous thymidine kinase but 5-fluorodeoxyuridine inhibits growth when exogenous thymidine kinase is expressed. Unexpectedly, we found that 5-fluorodeoxyuridine causes S phase arrest even without thymidine kinase expression. DNA damage checkpoint proteins such as the 9-1-1 complex were required for viability in the presence of 5-fluorodeoxyuridine. We also found that strains with circular chromosomes, due to loss of pot1+, which have higher levels of replication stress, were more sensitive to loss of the 9-1-1 complex in the presence of 5-fluorodeoxyuridine. Thus, our results suggest that strains carrying circular chromosomes exhibit a greater dependence on DNA damage checkpoints to ensure viability in the presence of 5-fluorodeoxyuridine compared to stains that have linear chromosomes.

  12. Fission yeast strains with circular chromosomes require the 9-1-1 checkpoint complex for the viability in response to the anti-cancer drug 5-fluorodeoxyuridine.

    Science.gov (United States)

    Shamim, Hossain Mohammad; Minami, Yukako; Tanaka, Daiki; Ukimori, Shinobu; Murray, Johanne M; Ueno, Masaru

    2017-01-01

    Thymidine kinase converts 5-fluorodeoxyuridine to 5-fluorodeoxyuridine monophosphate, which causes disruption of deoxynucleotide triphosphate ratios. The fission yeast Schizosaccharomyces pombe does not express endogenous thymidine kinase but 5-fluorodeoxyuridine inhibits growth when exogenous thymidine kinase is expressed. Unexpectedly, we found that 5-fluorodeoxyuridine causes S phase arrest even without thymidine kinase expression. DNA damage checkpoint proteins such as the 9-1-1 complex were required for viability in the presence of 5-fluorodeoxyuridine. We also found that strains with circular chromosomes, due to loss of pot1+, which have higher levels of replication stress, were more sensitive to loss of the 9-1-1 complex in the presence of 5-fluorodeoxyuridine. Thus, our results suggest that strains carrying circular chromosomes exhibit a greater dependence on DNA damage checkpoints to ensure viability in the presence of 5-fluorodeoxyuridine compared to stains that have linear chromosomes.

  13. Study of the G2/M cell cycle checkpoint in irradiated mammary epithelial cells overexpressing Cul-4A gene

    International Nuclear Information System (INIS)

    Gupta, Anu; Yang, L.-X.; Chen, L.-C.

    2002-01-01

    Purpose: Members of the cullin gene family are known to be involved in cell cycle control. One of the cullin genes, Cul-4A, is amplified and overexpressed in breast cancer cells. This study investigates the effect of Cul-4A overexpression upon G2/M cell cycle checkpoint after DNA damage induced by either ionizing or nonionizing radiation. Methods and Materials: The normal mammary epithelial cell line MCF10A was stably transfected with full-length Cul-4A cDNA. Independent clones of MCF10A cells that overexpress Cul-4A proteins were selected and treated with either 8 Gy of ionizing radiation or 7 J/M 2 of UV radiation. The profile of cell cycle progression and the accumulation of several cell cycle proteins were analyzed. Results: We found that overexpression of Cul-4A in MCF10A cells abrogated the G2/M cell cycle checkpoint in response to DNA damage induced by ionizing irradiation, but not to DNA damage induced by nonionizing radiation. Analysis of cell cycle proteins showed that after ionizing irradiation, p53 accumulated in the mock-transfected MCF10A cells, but not in the Cul-4A transfectants. Conclusion: Our results suggest a role for Cul-4A in tumorigenesis and/or tumor progression, possibly through disruption of cell cycle control

  14. G2 Checkpoint Responses in Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    Britt, Anne [Univ. of California, Davis, CA (United States)

    2013-03-18

    This project focused on the mechanism and biological significance of the G2 arrest response to replication stress in plants. We employed both forward and reverse genetic approaches to identify genes required for this response. A total of 3 different postdocs, 5 undergraduates, and 2 graduate students participated in the project. We identified several genes required for damage response in plants, including homologs of genes previously identified in animals (ATM and ATR), novel, a plant-specific genes (SOG1) and a gene known in animals but previously thought to be missing from the Arabidopsis genome (ATRIP). We characterized the transcriptome of gamma-irradiated plants, and found that plants, unlike animals, express a robust transcriptional response to damage, involving genes that regulate the cell cycle and DNA metabolism. This response requires both ATM and the transcription factor SOG1. We found that both ATM and ATR play a role in meiosis in plants. We also found that plants have a cell-type-specific programmed cell death response to ionizing radiation and UV light, and that this response requires ATR, ATM, and SOG1. These results were published in a series of 5 papers.

  15. Akt Kinase-Mediated Checkpoint of cGAS DNA Sensing Pathway

    Directory of Open Access Journals (Sweden)

    Gil Ju Seo

    2015-10-01

    Full Text Available Upon DNA stimulation, cyclic GMP-AMP synthase (cGAS synthesizes the second messenger cyclic GMP-AMP (cGAMP that binds to the STING, triggering antiviral interferon-β (IFN-β production. However, it has remained undetermined how hosts regulate cGAS enzymatic activity after the resolution of DNA immunogen. Here, we show that Akt kinase plays a negative role in cGAS-mediated anti-viral immune response. Akt phosphorylated the S291 or S305 residue of the enzymatic domain of mouse or human cGAS, respectively, and this phosphorylation robustly suppressed its enzymatic activity. Consequently, expression of activated Akt led to the reduction of cGAMP and IFN-β production and the increase of herpes simplex virus 1 replication, whereas treatment with Akt inhibitor augmented cGAS-mediated IFN-β production. Furthermore, expression of the phosphorylation-resistant cGAS S291A mutant enhanced IFN-β production upon DNA stimulation, HSV-1 infection, and vaccinia virus infection. Our study identifies an Akt kinase-mediated checkpoint to fine-tune hosts’ immune responses to DNA stimulation.

  16. Akt Kinase-Mediated Checkpoint of cGAS DNA Sensing Pathway.

    Science.gov (United States)

    Seo, Gil Ju; Yang, Aerin; Tan, Brandon; Kim, Sungyoon; Liang, Qiming; Choi, Younho; Yuan, Weiming; Feng, Pinghui; Park, Hee-Sung; Jung, Jae U

    2015-10-13

    Upon DNA stimulation, cyclic GMP-AMP synthase (cGAS) synthesizes the second messenger cyclic GMP-AMP (cGAMP) that binds to the STING, triggering antiviral interferon-β (IFN-β) production. However, it has remained undetermined how hosts regulate cGAS enzymatic activity after the resolution of DNA immunogen. Here, we show that Akt kinase plays a negative role in cGAS-mediated anti-viral immune response. Akt phosphorylated the S291 or S305 residue of the enzymatic domain of mouse or human cGAS, respectively, and this phosphorylation robustly suppressed its enzymatic activity. Consequently, expression of activated Akt led to the reduction of cGAMP and IFN-β production and the increase of herpes simplex virus 1 replication, whereas treatment with Akt inhibitor augmented cGAS-mediated IFN-β production. Furthermore, expression of the phosphorylation-resistant cGAS S291A mutant enhanced IFN-β production upon DNA stimulation, HSV-1 infection, and vaccinia virus infection. Our study identifies an Akt kinase-mediated checkpoint to fine-tune hosts' immune responses to DNA stimulation. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  17. Characterization of a cancer cell line that expresses a splicing variant form of 53BP1: Separation of checkpoint and repair functions in 53BP1

    International Nuclear Information System (INIS)

    Iwabuchi, Kuniyoshi; Matsui, Tadashi; Hashimoto, Mitsumasa; Matsumoto, Yoshihisa; Kurihara, Takayuki; Date, Takayasu

    2008-01-01

    53BP1 plays important roles in checkpoint signaling and repair for DNA double-strand breaks. We found that a colon cancer cell line, SW48, expressed a splicing variant form of 53BP1, which lacks the residues corresponding to exons 10 and 11. Activation of ATM and phosphorylation of ATM and ATR targets occurred in SW48 cells in response to X-irradiation, and these X-ray-induced responses were not enhanced by expression of full-length 53BP1 in SW48 cells, indicating that this splicing variant fully activates the major checkpoint signaling in SW48 cells. In contrast, the expression of full-length 53BP1 in SW48 cells promoted the repair of X-ray-induced DNA damage, evidenced by faster disappearance of X-ray-induced γ-H2AX foci, a marker for DNA damage, and less residual chromosomal aberrations after X-irradiation. We conclude that the two major roles of 53BP1, the checkpoint signaling and repair for DNA damage, can be functionally separated

  18. Covering Resilience: A Recent Development for Binomial Checkpointing

    Energy Technology Data Exchange (ETDEWEB)

    Walther, Andrea; Narayanan, Sri Hari Krishna

    2016-09-12

    In terms of computing time, adjoint methods offer a very attractive alternative to compute gradient information, required, e.g., for optimization purposes. However, together with this very favorable temporal complexity result comes a memory requirement that is in essence proportional with the operation count of the underlying function, e.g., if algorithmic differentiation is used to provide the adjoints. For this reason, checkpointing approaches in many variants have become popular. This paper analyzes an extension of the so-called binomial approach to cover also possible failures of the computing systems. Such a measure of precaution is of special interest for massive parallel simulations and adjoint calculations where the mean time between failure of the large scale computing system is smaller than the time needed to complete the calculation of the adjoint information. We describe the extensions of standard checkpointing approaches required for such resilience, provide a corresponding implementation and discuss first numerical results.

  19. Implementing forward recovery using checkpointing in distributed systems

    Science.gov (United States)

    Long, Junsheng; Fuchs, W. K.; Abraham, Jacob A.

    1991-01-01

    The paper describes the implementation of a forward recovery scheme using checkpoints and replicated tasks. The implementation is based on the concept of lookahead execution and rollback validation. In the experiment, two tasks are selected for the normal execution and one for rollback validation. It is shown that the recovery strategy has nearly error-free execution time and an average redundancy lower than TMR.

  20. Analysis of Drug Development Paradigms for Immune Checkpoint Inhibitors.

    Science.gov (United States)

    Jardim, Denis L; de Melo Gagliato, Débora; Giles, Francis J; Kurzrock, Razelle

    2018-04-15

    Immune checkpoint inhibitors have unique toxicities and response kinetics compared with cytotoxic and gene-targeted anticancer agents. We investigated the impact of innovative/accelerated immunotherapy drug development/approval models on the accuracy of safety and efficacy assessments by searching the FDA website. Initial phase I trials for each agent were reviewed and safety and efficacy data compared with that found in later trials leading to regulatory approvals of the same agents. As of June 2017, the FDA approved six checkpoint inhibitors for a variety of cancer types. All checkpoint inhibitors received a priority review status and access to at least two additional FDA special access programs, more often breakthrough therapy designation and accelerated approval. Median clinical development time (investigational new drug application to approval) was 60.77 months [avelumab had the shortest timeline (52.33 months)]. Response rates during early phase I trials (median = 16%) are higher than for phase I trials of other agents (with the exception of gene-targeted agents tested with a biomarker). Doses approved were usually not identical to doses recommended on phase I trials. Approximately 50% of types of immune-related and 43% of types of clinically relevant toxicities from later trials were identified in early-phase trials. Even so, treatment-related mortality remains exceedingly low in later studies (0.33% of patients). In conclusion, efficacy and safety of immune checkpoint inhibitors appear to be reasonably predicted from the dose-finding portion of phase I trials, indicating that the fast-track development of these agents is safe and justified. Clin Cancer Res; 24(8); 1785-94. ©2017 AACR . ©2017 American Association for Cancer Research.

  1. Spindle assembly checkpoint acquisition at the mid-blastula transition.

    Directory of Open Access Journals (Sweden)

    Maomao Zhang

    Full Text Available The spindle assembly checkpoint (SAC maintains the fidelity of chromosome segregation during mitosis. Nonpathogenic cells lacking the SAC are typically only found in cleavage stage metazoan embryos, which do not acquire functional checkpoints until the mid-blastula transition (MBT. It is unclear how proper SAC function is acquired at the MBT, though several models exist. First, SAC acquisition could rely on transcriptional activity, which increases dramatically at the MBT. Embryogenesis prior to the MBT relies primarily on maternally loaded transcripts, and if SAC signaling components are not maternally supplied, the SAC would depend on zygotic transcription at the MBT. Second, checkpoint acquisition could depend on the Chk1 kinase, which is activated at the MBT to elongate cell cycles and is required for the SAC in somatic cells. Third, SAC function could depend on a threshold nuclear to cytoplasmic (N:C ratio, which increases during pre-MBT cleavage cycles and dictates several MBT events like zygotic transcription and cell cycle remodeling. Finally, the SAC could by regulated by a timer mechanism that coincides with other MBT events but is independent of them. Using zebrafish embryos we show that SAC acquisition at the MBT is independent of zygotic transcription, indicating that the checkpoint program is maternally supplied. Additionally, by precociously lengthening cleavage cycles with exogenous Chk1 activity, we show that cell cycle lengthening and Chk1 activity are not sufficient for SAC acquisition. Furthermore, we find that SAC acquisition can be uncoupled from the N:C ratio. Together, our findings indicate that SAC acquisition is regulated by a maternally programmed developmental timer.

  2. Radiation-induced apoptosis and cell cycle checkpoints in human colorectal tumour cell lines

    International Nuclear Information System (INIS)

    Playle, L.C.

    2001-03-01

    The p53 tumour suppressor gene is mutated in 75% of colorectal carcinomas and is critical for DNA damage-induced G1 cell cycle arrest. Data presented in this thesis demonstrate that after treatment with Ionizing Radiation (IR), colorectal tumour cell lines with mutant p53 are unable to arrest at G1 and undergo cell cycle arrest at G2. The staurosporine derivative, UCN-01, was shown to abrogate the IR-induced G2 checkpoint in colorectal tumour cell lines. Furthermore, in some cell lines, abrogation of the G2 checkpoint was associated with radiosensitisation. Data presented in this study demonstrate that 2 out of 5 cell lines with mutant p53 were sensitised to IR by UCN-01. In order to determine whether radiosensitisation correlated with lack of functional p53, transfected derivatives of an adenoma-derived cell line were studied, in which endogenous wild type p53 was disrupted by expression of a dominant negative p53 mutant protein (and with a vector control). In both these cell lines UCN-01 abrogated the G2 arrest however this was not associated with radiosensitisation, indicating that radiosensitisation is a cell type-specific phenomenon. Although 2 colorectal carcinoma cell lines, with mutant p53, were sensitised to IR by UCN-01, the mechanisms of p53-independent IR-induced apoptosis in the colon are essentially unknown. The mitogen-activated protein kinase (MAPK) pathways (that is the JNK, p38 and ERK pathways) have been implicated in apoptosis in a range of cell systems and in IR-induced apoptosis in some cell types. Data presented in this study show that, although the MAPKs can be activated by the known activator anisomycin, there is no evidence of a role for MAPKs in IR-induced apoptosis in colorectal tumour cell lines, regardless of p53 status. In summary, some colorectal tumour cell lines with mutant p53 can be sensitised to IR-induced cell death by G2 checkpoint abrogation and this may be an important treatment strategy, however mechanisms of IR-induced p53

  3. Disruption of murine mp29/Syf2/Ntc31 gene results in embryonic lethality with aberrant checkpoint response.

    Directory of Open Access Journals (Sweden)

    Chia-Hsin Chen

    Full Text Available Human p29 is a putative component of spliceosomes, but its role in pre-mRNA is elusive. By siRNA knockdown and stable overexpression, we demonstrated that human p29 is involved in DNA damage response and Fanconi anemia pathway in cultured cells. In this study, we generated p29 knockout mice (mp29(GT/GT using the mp29 gene trap embryonic stem cells to study the role of mp29 in DNA damage response in vivo. Interruption of mp29 at both alleles resulted in embryonic lethality. Embryonic abnormality occurred as early as E6.5 in mp29(GT/GT mice accompanied with decreased mRNA levels of α-tubulin and Chk1. The reduction of α-tubulin and Chk1 mRNAs is likely due to an impaired post-transcriptional event. An aberrant G2/M checkpoint was found in mp29 gene trap embryos when exposed to aphidicolin and UV light. This embryonic lethality was rescued by crossing with mp29 transgenic mice. Additionally, the knockdown of zfp29 in zebrafish resulted in embryonic death at 72 hours of development postfertilization (hpf. A lower level of acetylated α-tubulin was also observed in zfp29 morphants. Together, these results illustrate an indispensable role of mp29 in DNA checkpoint response during embryonic development.

  4. Sobriety checkpoints in Thailand: a review of effectiveness and developments over time.

    Science.gov (United States)

    Ditsuwan, Vallop; Veerman, J Lennert; Bertram, Melanie; Vos, Theo

    2015-03-01

    This review describes the legal basis for and implementation of sobriety checkpoints in Thailand and identifies factors that influenced their historical development and effectiveness. The first alcohol and traffic injury control law in Thailand was implemented in 1934. The 0.05 g/100 mL blood alcohol concentration limit was set in 1994. Currently, 3 types of sobriety checkpoints are used: general police checkpoints, selective breath testing, and special event sobriety checkpoints. The authors found few reports on the strategies, frequencies, and outcomes for any of these types of checkpoints, despite Thailand having devoted many resources to their implementation. In Thailand and other low-middle income countries, it is necessary to address the country-specific barriers to successful enforcement (including political and logistical issues, lack of equipment, and absence of other supportive alcohol harm reduction measures) before sobriety checkpoints can be expected to be as effective as reported in high-income countries. © 2011 APJPH.

  5. Emodnet Med Sea Check-Point - Indicators for decision- maker

    Science.gov (United States)

    Besnard, Sophie; Claverie, Vincent; Blanc, Frédérique

    2015-04-01

    The Emodnet Checkpoint projects aim is to assess the cost-effectiveness, reliability and utility of the existing monitoring at the sea basin level. This involves the development of monitoring system indicators and a GIS Platform to perform the assessment and make it available. Assessment or production of Check-Point information is made by developing targeted products based on the monitoring data and determining whether the products are meeting the needs of industry and public authorities. Check-point users are the research community, the 'institutional' policy makers for IMP and MSFD implementation, the 'intermediate users', i.e., users capable to understand basic raw data but that benefit from seeing the Checkpoint targeted products and the assessment of the fitness for purpose. We define assessment criteria aimed to characterize/depict the input datasets in terms of 3 territories capable to show performance and gaps of the present monitoring system, appropriateness, availability and fitness for purpose. • Appropriateness: What is made available to users? What motivate/decide them to select this observation rather than this one. • Availability: How this is made available to the user? Place to understand the readiness and service performance of the EU infrastructure • Fitness for use / fitness for purpose: Ability for non-expert user to appreciate the data exploitability (feedback on efficiency & reliability of marine data) For each territory (appropriateness, Availability and Fitness for purpose / for use), we define several indicators. For example, for Availability we define Visibility, Accessibility and Performance. And Visibility is itself defined by "Easily found" and "EU service". So these indicators can be classified according to their territory and sub-territory as seen above, but also according to the complexity to build them. Indicators are built from raw descriptors in 3 stages:  Stage 1: to give a neutral and basic status directly computed from

  6. The DNA replication checkpoint directly regulates MBF-dependent G1/S transcription.

    Science.gov (United States)

    Dutta, Chaitali; Patel, Prasanta K; Rosebrock, Adam; Oliva, Anna; Leatherwood, Janet; Rhind, Nicholas

    2008-10-01

    The DNA replication checkpoint transcriptionally upregulates genes that allow cells to adapt to and survive replication stress. Our results show that, in the fission yeast Schizosaccharomyces pombe, the replication checkpoint regulates the entire G(1)/S transcriptional program by directly regulating MBF, the G(1)/S transcription factor. Instead of initiating a checkpoint-specific transcriptional program, the replication checkpoint targets MBF to maintain the normal G(1)/S transcriptional program during replication stress. We propose a mechanism for this regulation, based on in vitro phosphorylation of the Cdc10 subunit of MBF by the Cds1 replication-checkpoint kinase. Replacement of two potential phosphorylation sites with phosphomimetic amino acids suffices to promote the checkpoint transcriptional program, suggesting that Cds1 phosphorylation directly regulates MBF-dependent transcription. The conservation of MBF between fission and budding yeast, and recent results implicating MBF as a target of the budding yeast replication checkpoint, suggests that checkpoint regulation of the MBF transcription factor is a conserved strategy for coping with replication stress. Furthermore, the structural and regulatory similarity between MBF and E2F, the metazoan G(1)/S transcription factor, suggests that this checkpoint mechanism may be broadly conserved among eukaryotes.

  7. Action-oriented use of ergonomic checkpoints for healthy work design in different settings.

    Science.gov (United States)

    Kogi, Kazutaka

    2007-12-01

    Recent experiences in the action-oriented use of ergonomic checkpoints in different work settings are reviewed. The purpose is to know what features are useful for healthy work design adjusted to each local situation. Based on the review results, common features of ergonomic checkpoints used in participatory training programs for improving workplace conditions in small enterprises, construction sites, home work and agriculture in industrially developing countries in Asia are discussed. These checkpoints generally compile practical improvement options in a broad range of technical areas, such as materials handling, workstation design, physical environment and work organization. Usually, "action checklists" comprising the tiles of the checkpoints are used together. A clear focus is placed on readily applicable low-cost options. Three common features of these various checkpoints appear to be important. First, the checkpoints represent typical good practices in multiple areas. Second, each how-to section of these checkpoints presents simple improvements reflecting basic ergonomic principles. Examples of these principles include easy reach, fewer and faster transport, elbow-level work, coded displays, isolated or screened hazards and shared teamwork. Third, the illustrated checkpoints accompanied by corresponding checklists are used as group work tools in short-term training courses. Many practical improvements achieved are displayed in websites for inter-country work improvement networks. It is suggested to promote the use of locally adjusted checkpoints in various forms of participatory action-oriented training in small-scale workplaces and in agriculture particularly in industrially developing countries.

  8. The DNA Replication Checkpoint Directly Regulates MBF-Dependent G1/S Transcription▿

    Science.gov (United States)

    Dutta, Chaitali; Patel, Prasanta K.; Rosebrock, Adam; Oliva, Anna; Leatherwood, Janet; Rhind, Nicholas

    2008-01-01

    The DNA replication checkpoint transcriptionally upregulates genes that allow cells to adapt to and survive replication stress. Our results show that, in the fission yeast Schizosaccharomyces pombe, the replication checkpoint regulates the entire G1/S transcriptional program by directly regulating MBF, the G1/S transcription factor. Instead of initiating a checkpoint-specific transcriptional program, the replication checkpoint targets MBF to maintain the normal G1/S transcriptional program during replication stress. We propose a mechanism for this regulation, based on in vitro phosphorylation of the Cdc10 subunit of MBF by the Cds1 replication-checkpoint kinase. Replacement of two potential phosphorylation sites with phosphomimetic amino acids suffices to promote the checkpoint transcriptional program, suggesting that Cds1 phosphorylation directly regulates MBF-dependent transcription. The conservation of MBF between fission and budding yeast, and recent results implicating MBF as a target of the budding yeast replication checkpoint, suggests that checkpoint regulation of the MBF transcription factor is a conserved strategy for coping with replication stress. Furthermore, the structural and regulatory similarity between MBF and E2F, the metazoan G1/S transcription factor, suggests that this checkpoint mechanism may be broadly conserved among eukaryotes. PMID:18662996

  9. Genetic Control of the Trigger for the G2/M Checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Hall, Eric J. [Columbia University; Smilenov, Lubomir B. [Columbia University; Young, Erik F. [Columbia University

    2013-10-01

    The work undertaken in this project addressed two seminal areas of low dose radiation biology that are poorly understood and controversial. These areas are the challenge to the linear-no-threshold (LNT) paradigm at low doses of radiation and, the fundamental elements of radiation bystander effect biology Genetic contributions to low dose checkpoint engagement: The LNT paradigm is an extrapolation of known, measured cancer induction endpoints. Importantly, data for lower doses is often not available. Debatably, radiation protection standards have been introduced which are prudently contingent on the adherence of cancer risk to the established trend seen at higher doses. Intriguing findings from other labs have hinted at separate DNA damage response programs that engage at low or high levels of radiation. Individual radiation sensitivity commensurate with hemizygosity for a radiation sensitivity gene has been estimated at 1-2% in the U.S.. Careful interrogation of the DNA damage response at low doses of radiation became important and served as the basis for this grant. Several genes were tested in combinations to determine if combined haploinsufficiency for multiple radiosensitizing genes could render a cell more sensitive to lower levels of acute radiation exposure. We measured a classical radiation response endpoint, cell cycle arrest prior to mitosis. Mouse embryo fibroblasts were used and provided a uniform, rapidly dividing and genetically manipulable population of study. Our system did not report checkpoint engagement at acute doses of gamma rays below 100 mGy. The system did report checkpoint engagement reproducibly at 500 mGy establishing a threshold for activation between 100 and 500 mGy. Engagement of the checkpoint was ablated in cells nullizygous for ATM but was otherwise unperturbed in cells combinatorially haploinsufficient for ATM and Rad9, ATM and PTEN or PTEN and Rad9. Taken together, these experiments tell us that, in a sensitive fibroblast culture

  10. The Neurospora crassa UVS-3 epistasis group encodes homologues of the ATR/ATRIP checkpoint control system.

    Science.gov (United States)

    Kazama, Yusuke; Ishii, Chizu; Schroeder, Alice L; Shimada, Hisao; Wakabayashi, Michiyoshi; Inoue, Hirokazu

    2008-02-01

    The mutagen sensitive uvs-3 and mus-9 mutants of Neurospora show mutagen and hydroxyurea sensitivity, mutator effects and duplication instability typical of recombination repair and DNA damage checkpoint defective mutants. To determine the nature of these genes we used cosmids from a genomic library to clone the uvs-3 gene by complementation for MMS sensitivity. Mutation induction by transposon insertion and RIP defined the coding sequence. RFLP analysis confirmed that this sequence maps in the area of uvs-3 at the left telomere of LG IV. Analysis of the cDNA showed that the UVS-3 protein contains an ORF of 969 amino acids with one intron. It is homologous to UvsD of Aspergillus nidulans, a member of the ATRIP family of checkpoint proteins. It retains the N' terminal coiled-coil motif followed by four basic amino acids typical of these proteins and shows the highest homology in this region. The uvsD cDNA partially complements the defects of the uvs-3 mutation. The uvs-3 mutant shows a higher level of micronuclei in conidia and failure to halt germination and nuclear division in the presence of hydroxyurea than wild type, suggesting checkpoint defects. ATRIP proteins bind tightly to ATR PI-3 kinase (phosphatidylinositol 3-kinase) proteins. Therefore, we searched the Neurospora genome sequence for homologues of the Aspergillus nidulans ATR, UvsB. A uvsB homologous sequence was present in the right arm of chromosome I where the mus-9 gene maps. A cosmid containing this genomic DNA complemented the mus-9 mutation. The putative MUS-9 protein is 2484 amino acids long with eight introns. Homology is especially high in the C-terminal 350 amino acids that correspond to the PI-3 kinase domain. In wild type a low level of constitutive mRNA is present for both genes. It is transiently induced upon UV exposure.

  11. Multiple Duties for Spindle Assembly Checkpoint Kinases in Meiosis

    Science.gov (United States)

    Marston, Adele L.; Wassmann, Katja

    2017-01-01

    Cell division in mitosis and meiosis is governed by evolutionary highly conserved protein kinases and phosphatases, controlling the timely execution of key events such as nuclear envelope breakdown, spindle assembly, chromosome attachment to the spindle and chromosome segregation, and cell cycle exit. In mitosis, the spindle assembly checkpoint (SAC) controls the proper attachment to and alignment of chromosomes on the spindle. The SAC detects errors and induces a cell cycle arrest in metaphase, preventing chromatid separation. Once all chromosomes are properly attached, the SAC-dependent arrest is relieved and chromatids separate evenly into daughter cells. The signaling cascade leading to checkpoint arrest depends on several protein kinases that are conserved from yeast to man. In meiosis, haploid cells containing new genetic combinations are generated from a diploid cell through two specialized cell divisions. Though apparently less robust, SAC control also exists in meiosis. Recently, it has emerged that SAC kinases have additional roles in executing accurate chromosome segregation during the meiotic divisions. Here, we summarize the main differences between mitotic and meiotic cell divisions, and explain why meiotic divisions pose special challenges for correct chromosome segregation. The less-known meiotic roles of the SAC kinases are described, with a focus on two model systems: yeast and mouse oocytes. The meiotic roles of the canonical checkpoint kinases Bub1, Mps1, the pseudokinase BubR1 (Mad3), and Aurora B and C (Ipl1) will be discussed. Insights into the molecular signaling pathways that bring about the special chromosome segregation pattern during meiosis will help us understand why human oocytes are so frequently aneuploid. PMID:29322045

  12. Multiple Duties for Spindle Assembly Checkpoint Kinases in Meiosis

    Directory of Open Access Journals (Sweden)

    Adele L. Marston

    2017-12-01

    Full Text Available Cell division in mitosis and meiosis is governed by evolutionary highly conserved protein kinases and phosphatases, controlling the timely execution of key events such as nuclear envelope breakdown, spindle assembly, chromosome attachment to the spindle and chromosome segregation, and cell cycle exit. In mitosis, the spindle assembly checkpoint (SAC controls the proper attachment to and alignment of chromosomes on the spindle. The SAC detects errors and induces a cell cycle arrest in metaphase, preventing chromatid separation. Once all chromosomes are properly attached, the SAC-dependent arrest is relieved and chromatids separate evenly into daughter cells. The signaling cascade leading to checkpoint arrest depends on several protein kinases that are conserved from yeast to man. In meiosis, haploid cells containing new genetic combinations are generated from a diploid cell through two specialized cell divisions. Though apparently less robust, SAC control also exists in meiosis. Recently, it has emerged that SAC kinases have additional roles in executing accurate chromosome segregation during the meiotic divisions. Here, we summarize the main differences between mitotic and meiotic cell divisions, and explain why meiotic divisions pose special challenges for correct chromosome segregation. The less-known meiotic roles of the SAC kinases are described, with a focus on two model systems: yeast and mouse oocytes. The meiotic roles of the canonical checkpoint kinases Bub1, Mps1, the pseudokinase BubR1 (Mad3, and Aurora B and C (Ipl1 will be discussed. Insights into the molecular signaling pathways that bring about the special chromosome segregation pattern during meiosis will help us understand why human oocytes are so frequently aneuploid.

  13. Systematic Analysis of the DNA Damage Response Network in Telomere Defective Budding Yeast

    Directory of Open Access Journals (Sweden)

    Eva-Maria Holstein

    2017-07-01

    Full Text Available Functional telomeres are critically important to eukaryotic genetic stability. Scores of proteins and pathways are known to affect telomere function. Here, we report a series of related genome-wide genetic interaction screens performed on budding yeast cells with acute or chronic telomere defects. Genetic interactions were examined in cells defective in Cdc13 and Stn1, affecting two components of CST, a single stranded DNA (ssDNA binding complex that binds telomeric DNA. For comparison, genetic interactions were also examined in cells with defects in Rfa3, affecting the major ssDNA binding protein, RPA, which has overlapping functions with CST at telomeres. In more complex experiments, genetic interactions were measured in cells lacking EXO1 or RAD9, affecting different aspects of the DNA damage response, and containing a cdc13-1 induced telomere defect. Comparing fitness profiles across these data sets helps build a picture of the specific responses to different types of dysfunctional telomeres. The experiments show that each context reveals different genetic interactions, consistent with the idea that each genetic defect causes distinct molecular defects. To help others engage with the large volumes of data, the data are made available via two interactive web-based tools: Profilyzer and DIXY. One particularly striking genetic interaction observed was that the chk1∆ mutation improved fitness of cdc13-1 exo1∆ cells more than other checkpoint mutations (ddc1∆, rad9∆, rad17∆, and rad24∆, whereas, in cdc13-1 cells, the effects of all checkpoint mutations were similar. We show that this can be explained by Chk1 stimulating resection—a new function for Chk1 in the eukaryotic DNA damage response network.

  14. Predictors of responses to immune checkpoint blockade in advanced melanoma

    DEFF Research Database (Denmark)

    Jacquelot, N; Roberti, M P; Enot, D P

    2017-01-01

    Immune checkpoint blockers (ICB) have become pivotal therapies in the clinical armamentarium against metastatic melanoma (MMel). Given the frequency of immune related adverse events and increasing use of ICB, predictors of response to CTLA-4 and/or PD-1 blockade represent unmet clinical needs....... Using a systems biology-based approach to an assessment of 779 paired blood and tumor markers in 37 stage III MMel patients, we analyzed association between blood immune parameters and the functional immune reactivity of tumor-infiltrating cells after ex vivo exposure to ICB. Based on this assay, we...

  15. Central Tolerance Blockade to Augment Checkpoint Immunotherapy in Melanoma

    Science.gov (United States)

    2017-09-01

    expressed at higher levels in medullary thymic epithelial cells (mTECs) than in cortical thymic epithelial cells (cTECs), addition of anti-RANKL antibody...relative Aire expression by quantitative RT-PCR in cultured thymic tissue . 12-15 90% Subtask 3. Culture human thymus sections with OPG-Fc or vehicle...PCR in cultured thymic tissue . 15-18 90% ACURO approval 13-14 100% Major Task 1. Effect of concurrent anti-RANKL and checkpoint inhibitor

  16. Conformation-specific anti-Mad2 monoclonal antibodies for the dissection of checkpoint signaling

    DEFF Research Database (Denmark)

    Sedgwick, Garry G; Larsen, Marie Sofie Yoo; Lischetti, Tiziana

    2016-01-01

    The spindle assembly checkpoint (SAC) ensures accurate chromosome segregation during mitosis by delaying the activation of the anaphase-promoting complex/cyclosome (APC/C) in response to unattached kinetochores. The Mad2 protein is essential for a functional checkpoint because it binds directly t...

  17. Space Reclamation for Uncoordinated Checkpointing in Message-Passing Systems. Ph.D. Thesis

    Science.gov (United States)

    Wang, Yi-Min

    1993-01-01

    Checkpointing and rollback recovery are techniques that can provide efficient recovery from transient process failures. In a message-passing system, the rollback of a message sender may cause the rollback of the corresponding receiver, and the system needs to roll back to a consistent set of checkpoints called recovery line. If the processes are allowed to take uncoordinated checkpoints, the above rollback propagation may result in the domino effect which prevents recovery line progression. Traditionally, only obsolete checkpoints before the global recovery line can be discarded, and the necessary and sufficient condition for identifying all garbage checkpoints has remained an open problem. A necessary and sufficient condition for achieving optimal garbage collection is derived and it is proved that the number of useful checkpoints is bounded by N(N+1)/2, where N is the number of processes. The approach is based on the maximum-sized antichain model of consistent global checkpoints and the technique of recovery line transformation and decomposition. It is also shown that, for systems requiring message logging to record in-transit messages, the same approach can be used to achieve optimal message log reclamation. As a final topic, a unifying framework is described by considering checkpoint coordination and exploiting piecewise determinism as mechanisms for bounding rollback propagation, and the applicability of the optimal garbage collection algorithm to domino-free recovery protocols is demonstrated.

  18. Rituximab does not reset defective early B cell tolerance checkpoints

    Science.gov (United States)

    Chamberlain, Nicolas; Massad, Christopher; Oe, Tyler; Cantaert, Tineke; Herold, Kevan C.; Meffre, Eric

    2015-01-01

    Type 1 diabetes (T1D) patients show abnormalities in early B cell tolerance checkpoints, resulting in the accumulation of large numbers of autoreactive B cells in their blood. Treatment with rituximab, an anti-CD20 mAb that depletes B cells, has been shown to preserve β cell function in T1D patients and improve other autoimmune diseases, including rheumatoid arthritis and multiple sclerosis. However, it remains largely unknown how anti–B cell therapy thwarts autoimmunity in these pathologies. Here, we analyzed the reactivity of Abs expressed by single, mature naive B cells from 4 patients with T1D before and 52 weeks after treatment to determine whether rituximab resets early B cell tolerance checkpoints. We found that anti–B cell therapy did not alter the frequencies of autoreactive and polyreactive B cells, which remained elevated in the blood of all patients after rituximab treatment. Moreover, the limited proliferative history of autoreactive B cells after treatment revealed that these clones were newly generated B cells and not self-reactive B cells that had escaped depletion and repopulated the periphery through homeostatic expansion. We conclude that anti–B cell therapy may provide a temporary dampening of autoimmune processes through B cell depletion. However, repletion with autoreactive B cells may explain the relapse that occurs in many autoimmune patients after anti–B cell therapy. PMID:26642366

  19. Immunotherapy targeting immune check-point(s) in brain metastases.

    Science.gov (United States)

    Di Giacomo, Anna Maria; Valente, Monica; Covre, Alessia; Danielli, Riccardo; Maio, Michele

    2017-08-01

    Immunotherapy with monoclonal antibodies (mAb) directed to different immune check-point(s) is showing a significant clinical impact in a growing number of human tumors of different histotype, both in terms of disease response and long-term survival patients. In this rapidly changing scenario, treatment of brain metastases remains an high unmeet medical need, and the efficacy of immunotherapy in these highly dismal clinical setting remains to be largely demonstrated. Nevertheless, up-coming observations are beginning to suggest a clinical potential of cancer immunotherapy also in brain metastases, regardless the underlying tumor histotype. These observations remain to be validated in larger clinical trials eventually designed also to address the efficacy of therapeutic mAb to immune check-point(s) within multimodality therapies for brain metastases. Noteworthy, the initial proofs of efficacy on immunotherapy in central nervous system metastases are already fostering clinical trials investigating its therapeutic potential also in primary brain tumors. We here review ongoing immunotherapeutic approaches to brain metastases and primary brain tumors, and the foreseeable strategies to overcome their main biologic hurdles and clinical challenges. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Avelumab: combining immune checkpoint inhibition and antibody-dependent cytotoxicity.

    Science.gov (United States)

    Hamilton, Gerhard; Rath, Barbara

    2017-04-01

    Immune checkpoint inhibition holds great promise for selected tumors. The human monoclonal antibody (mAB) avelumab is directed to programmed death ligand-1 (PD-L1) and is supposed to inhibit the immunosuppressive PD-L1/PD-1 interaction and, furthermore, effect antibody-dependent cytotoxicity (ADCC) lysis of tumor cells. Areas covered: This article presents an overview of the current means to activate the antitumor immune defense by targeting PD-1 or PD-L1 with mABs and their possible role in ADCC-mediated tumor cell elimination. Expert opinion: Avelumab contains a Fc region which can bind cognate receptors on immune effector cells and induce ADCC-mediated tumor cell lysis, in contrast to other mABs directed to PD-1/PD-L1 which lack the ability to trigger ADCC due to belonging to the IgG4 subclass or possessing a mutated Fc region. Preclinical and clinical data indicate that avelumab can be safely administered to cancer patients with a toxicity profile comparable to other mABs and without lysis of PD-L1-positive activated immune cells. This antibody yielded durable responses in a phase II trial in advanced Merkel cell carcinoma patients. Tumor cell lysis by avelumab prevents cells from resorting to alternative checkpoints as shown by targeting PD-1 and the upregulation of TIM-3.

  1. Cenp-meta is required for sustained spindle checkpoint

    Directory of Open Access Journals (Sweden)

    Thomas Rubin

    2014-05-01

    Full Text Available Cenp-E is a kinesin-like motor protein required for efficient end-on attachment of kinetochores to the spindle microtubules. Cenp-E immunodepletion in Xenopus mitotic extracts results in the loss of mitotic arrest and massive chromosome missegregation, whereas its depletion in mammalian cells leads to chromosome segregation defects despite the presence of a functional spindle assembly checkpoint (SAC. Cenp-meta has previously been reported to be the Drosophila homolog of vertebrate Cenp-E. In this study, we show that cenp-metaΔ mutant neuroblasts arrest in mitosis when treated with colchicine. cenp-metaΔ mutant cells display a mitotic delay. Yet, despite the persistence of the two checkpoint proteins Mad2 and BubR1 on unattached kinetochores, these cells eventually enter anaphase and give rise to highly aneuploid daughter cells. Indeed, we find that cenp-metaΔ mutant cells display a slow but continuous degradation of cyclin B, which eventually triggers the mitotic exit observed. Thus, our data provide evidence for a role of Cenp-meta in sustaining the SAC response.

  2. The tumor suppressor homolog in fission yeast, myh1{sup +}, displays a strong interaction with the checkpoint gene rad1{sup +}

    Energy Technology Data Exchange (ETDEWEB)

    Jansson, Kristina; Warringer, Jonas; Farewell, Anne [Department of Cell and Molecular Biology, Lundberg Laboratory, Goeteborg University, P.O. Box 462, Goeteborg SE-405 30 (Sweden); Park, Han-Oh [Bioneer Corporation, 49-3, Munpyeong-dong, Daedeok-gu, Daejon 306-220 (Korea, Republic of); Hoe, Kwang-Lae; Kim, Dong-Uk [Functional Genomics Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Yusong, Daejeon (Korea, Republic of); Hayles, Jacqueline [Cell Cycle Laboratory, Cancer Research UK, London Research Institute, 44 Lincoln' s Inn Fields, London WC2A 3PX (United Kingdom); Sunnerhagen, Per [Department of Cell and Molecular Biology, Lundberg Laboratory, Goeteborg University, P.O. Box 462, Goeteborg SE-405 30 (Sweden)], E-mail: per.sunnerhagen@cmb.gu.se

    2008-09-26

    The DNA glycosylase MutY is strongly conserved in evolution, and homologs are found in most eukaryotes and prokaryotes examined. This protein is implicated in repair of oxidative DNA damage, in particular adenine mispaired opposite 7,8-dihydro-8-oxoguanine. Previous investigations in Escherichia coli, fission yeast, and mammalian cells show an association of mutations in MutY homologs with a mutator phenotype and carcinogenesis. Eukaryotic MutY homologs physically associate with several proteins with a role in replication, DNA repair, and checkpoint signaling, specifically the trimeric 9-1-1 complex. In a genetic investigation of the fission yeast MutY homolog, myh1{sup +}, we show that the myh1 mutation confers a moderately increased UV sensitivity alone and in combination with mutations in several DNA repair genes. The myh1 rad1, and to a lesser degree myh1 rad9, double mutants display a synthetic interaction resulting in enhanced sensitivity to DNA damaging agents and hydroxyurea. UV irradiation of myh1 rad1 double mutants results in severe chromosome segregation defects and visible DNA fragmentation, and a failure to activate the checkpoint. Additionally, myh1 rad1 double mutants exhibit morphological defects in the absence of DNA damaging agents. We also found a moderate suppression of the slow growth and UV sensitivity of rhp51 mutants by the myh1 mutation. Our results implicate fission yeast Myh1 in repair of a wider range of DNA damage than previously thought, and functionally link it to the checkpoint pathway.

  3. Ultraviolet B, melanin and mitochondrial DNA: Photo-damage in human epidermal keratinocytes and melanocytes modulated by alpha-melanocyte-stimulating hormone [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Markus Böhm

    2016-05-01

    Full Text Available Alpha-melanocyte-stimulating hormone (alpha-MSH increases melanogenesis and protects from UV-induced DNA damage. However, its effect on mitochondrial DNA (mtDNA damage is unknown. We have addressed this issue in a pilot study using human epidermal keratinocytes and melanocytes incubated with alpha-MSH and irradiated with UVB. Real-time touchdown PCR was used to quantify total and deleted mtDNA. The deletion detected encompassed the common deletion but was more sensitive to detection. There were 4.4 times more mtDNA copies in keratinocytes than in melanocytes. Irradiation alone did not affect copy numbers. Alpha-MSH slightly increased copy numbers in both cell types in the absence of UVB and caused a similar small decrease in copy number with dose in both cell types. Deleted copies were nearly twice as frequent in keratinocytes as in melanocytes. Alpha-MSH reduced the frequency of deleted copies by half in keratinocytes but not in melanocytes. UVB dose dependently led to an increase in the deleted copy number in alpha-MSH-treated melanocytes. UVB irradiation had little effect on deleted copy number in alpha-MSH-treated keratinocytes. In summary, alpha-MSH enhances mtDNA damage in melanocytes presumably by increased melanogenesis, while α-MSH is protective in keratinocytes, the more so in the absence of irradiation.

  4. Mitotic catastrophe occurs in the absence of apoptosis in p53-null cells with a defective G1 checkpoint.

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    Michalis Fragkos

    Full Text Available Cell death occurring during mitosis, or mitotic catastrophe, often takes place in conjunction with apoptosis, but the conditions in which mitotic catastrophe may exhibit features of programmed cell death are still unclear. In the work presented here, we studied mitotic cell death by making use of a UV-inactivated parvovirus (adeno-associated virus; AAV that has been shown to induce a DNA damage response and subsequent death of p53-defective cells in mitosis, without affecting the integrity of the host genome. Osteosarcoma cells (U2OSp53DD that are deficient in p53 and lack the G1 cell cycle checkpoint respond to AAV infection through a transient G2 arrest. We found that the infected U2OSp53DD cells died through mitotic catastrophe with no signs of chromosome condensation or DNA fragmentation. Moreover, cell death was independent of caspases, apoptosis-inducing factor (AIF, autophagy and necroptosis. These findings were confirmed by time-lapse microscopy of cellular morphology following AAV infection. The assays used readily revealed apoptosis in other cell types when it was indeed occurring. Taken together the results indicate that in the absence of the G1 checkpoint, mitotic catastrophe occurs in these p53-null cells predominantly as a result of mechanical disruption induced by centrosome overduplication, and not as a consequence of a suicide signal.

  5. Checkpoint kinase 1-induced phosphorylation of O-linked β-N-acetylglucosamine transferase regulates the intermediate filament network during cytokinesis.

    Science.gov (United States)

    Li, Zhe; Li, Xueyan; Nai, Shanshan; Geng, Qizhi; Liao, Ji; Xu, Xingzhi; Li, Jing

    2017-12-01

    Checkpoint kinase 1 (Chk1) is a kinase instrumental for orchestrating DNA replication, DNA damage checkpoints, the spindle assembly checkpoint, and cytokinesis. Despite Chk1's pivotal role in multiple cellular processes, many of its substrates remain elusive. Here, we identified O- linked β- N -acetylglucosamine ( O -GlcNAc)-transferase (OGT) as one of Chk1's substrates. We found that Chk1 interacts with and phosphorylates OGT at Ser-20, which not only stabilizes OGT, but also is required for cytokinesis. Phospho-specific antibodies of OGT-pSer-20 exhibited specific signals at the midbody of the cell, consistent with midbody localization of OGT as reported previously. Moreover, phospho-deficient OGT (S20A) cells attenuated cellular O -GlcNAcylation levels and also reduced phosphorylation of Ser-71 in the cytoskeletal protein vimentin, a modification critical for severing vimentin filament during cytokinesis. Consequently, elongated vimentin bridges were observed in cells depleted of OGT via an si OGT- based approach. Lastly, expression of plasmids resistant to si OGT efficiently rescued the vimentin bridge phenotype, but the OGT-S20A rescue plasmids did not. Our results suggest a Chk1-OGT-vimentin pathway that regulates the intermediate filament network during cytokinesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Parallelization and checkpointing of GPU applications through program transformation

    Energy Technology Data Exchange (ETDEWEB)

    Solano-Quinde, Lizandro Damian [Iowa State Univ., Ames, IA (United States)

    2012-01-01

    GPUs have emerged as a powerful tool for accelerating general-purpose applications. The availability of programming languages that makes writing general-purpose applications for running on GPUs tractable have consolidated GPUs as an alternative for accelerating general purpose applications. Among the areas that have benefited from GPU acceleration are: signal and image processing, computational fluid dynamics, quantum chemistry, and, in general, the High Performance Computing (HPC) Industry. In order to continue to exploit higher levels of parallelism with GPUs, multi-GPU systems are gaining popularity. In this context, single-GPU applications are parallelized for running in multi-GPU systems. Furthermore, multi-GPU systems help to solve the GPU memory limitation for applications with large application memory footprint. Parallelizing single-GPU applications has been approached by libraries that distribute the workload at runtime, however, they impose execution overhead and are not portable. On the other hand, on traditional CPU systems, parallelization has been approached through application transformation at pre-compile time, which enhances the application to distribute the workload at application level and does not have the issues of library-based approaches. Hence, a parallelization scheme for GPU systems based on application transformation is needed. Like any computing engine of today, reliability is also a concern in GPUs. GPUs are vulnerable to transient and permanent failures. Current checkpoint/restart techniques are not suitable for systems with GPUs. Checkpointing for GPU systems present new and interesting challenges, primarily due to the natural differences imposed by the hardware design, the memory subsystem architecture, the massive number of threads, and the limited amount of synchronization among threads. Therefore, a checkpoint/restart technique suitable for GPU systems is needed. The goal of this work is to exploit higher levels of parallelism and

  7. Modulation of dendritic cell and T cell cross-talk during aging: The potential role of checkpoint inhibitory molecules.

    Science.gov (United States)

    Gardner, Joanne K; Mamotte, Cyril D S; Jackaman, Connie; Nelson, Delia J

    2017-09-01

    Dendritic cells (DCs) undergo continuous changes throughout life, and there is evidence that elderly DCs have a reduced capacity to stimulate T cells, which may contribute to impaired anti-tumour immune responses in elderly people with cancer. Changes in checkpoint inhibitory molecules/pathways during aging may be one mechanism that impairs the ability of elderly DCs to activate T cells. However, little is currently known regarding the combined effects of aging and cancer on DC and T cell inhibitory molecules/pathways. In this review, we discuss our current understanding of the influence of aging and cancer on key DC and T cell inhibitory molecules/pathways, the potential underlying cellular and molecular mechanisms contributing to their modulation, and the possibility of therapeutically targeting inhibitory molecules in elderly cancer patients. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. ALDH1A1 maintains ovarian cancer stem cell-like properties by altered regulation of cell cycle checkpoint and DNA repair network signaling.

    Directory of Open Access Journals (Sweden)

    Erhong Meng

    Full Text Available OBJECTIVE: Aldehyde dehydrogenase (ALDH expressing cells have been characterized as possessing stem cell-like properties. We evaluated ALDH+ ovarian cancer stem cell-like properties and their role in platinum resistance. METHODS: Isogenic ovarian cancer cell lines for platinum sensitivity (A2780 and platinum resistant (A2780/CP70 as well as ascites from ovarian cancer patients were analyzed for ALDH+ by flow cytometry to determine its association to platinum resistance, recurrence and survival. A stable shRNA knockdown model for ALDH1A1 was utilized to determine its effect on cancer stem cell-like properties, cell cycle checkpoints, and DNA repair mediators. RESULTS: ALDH status directly correlated to platinum resistance in primary ovarian cancer samples obtained from ascites. Patients with ALDHHIGH displayed significantly lower progression free survival than the patients with ALDHLOW cells (9 vs. 3 months, respectively p<0.01. ALDH1A1-knockdown significantly attenuated clonogenic potential, PARP-1 protein levels, and reversed inherent platinum resistance. ALDH1A1-knockdown resulted in dramatic decrease of KLF4 and p21 protein levels thereby leading to S and G2 phase accumulation of cells. Increases in S and G2 cells demonstrated increased expression of replication stress associated Fanconi Anemia DNA repair proteins (FANCD2, FANCJ and replication checkpoint (pS317 Chk1 were affected. ALDH1A1-knockdown induced DNA damage, evidenced by robust induction of γ-H2AX and BAX mediated apoptosis, with significant increases in BRCA1 expression, suggesting ALDH1A1-dependent regulation of cell cycle checkpoints and DNA repair networks in ovarian cancer stem-like cells. CONCLUSION: This data suggests that ovarian cancer cells expressing ALDH1A1 may maintain platinum resistance by altered regulation of cell cycle checkpoint and DNA repair network signaling.

  9. Inhibition of checkpoint kinase 1 sensitizes lung cancer brain metastases to radiotherapy

    International Nuclear Information System (INIS)

    Yang, Heekyoung; Yoon, Su Jin; Jin, Juyoun; Choi, Seung Ho; Seol, Ho Jun; Lee, Jung-Il

    2011-01-01

    Research highlights: → The most important therapeutic tool in brain metastasis is radiation therapy. → Radiosensitivity of cancer cells was enhanced with treatment of Chk1 inhibitor. → Depletion of Chk1 in cancer cells showed an enhancement of sensitivity to radiation. → Chk1 can be a good target for enhancement of radiosensitivity. -- Abstract: The most important therapeutic tool in brain metastasis is radiation therapy. However, resistance to radiation is a possible cause of recurrence or treatment failure. Recently, signal pathways about DNA damage checkpoints after irradiation have been noticed. We investigated the radiosensitivity can be enhanced with treatment of Chk1 inhibitor, AZD7762 in lung cancer cell lines and xenograft models of lung cancer brain metastasis. Clonogenic survival assays showed enhancement of radiosensitivity with AZD7762 after irradiation of various doses. AZD7762 increased ATR/ATM-mediated Chk1 phosphorylation and stabilized Cdc25A, suppressed cyclin A expression in lung cancer cell lines. In xenograft models of lung cancer (PC14PE6) brain metastasis, AZD7762 significantly prolonged the median survival time in response to radiation. Depletion of Chk1 using shRNA also showed an enhancement of sensitivity to radiation in PC14PE6 cells. The results of this study support that Chk1 can be a good target for enhancement of radiosensitivity.

  10. Adventures in Scaffold Morphing: Discovery of Fused Ring Heterocyclic Checkpoint Kinase 1 (CHK1) Inhibitors.

    Science.gov (United States)

    Yang, Bin; Vasbinder, Melissa M; Hird, Alexander W; Su, Qibin; Wang, Haixia; Yu, Yan; Toader, Dorin; Lyne, Paul D; Read, Jon A; Breed, Jason; Ioannidis, Stephanos; Deng, Chun; Grondine, Michael; DeGrace, Nancy; Whitston, David; Brassil, Patrick; Janetka, James W

    2018-02-08

    Checkpoint kinase 1 (CHK1) inhibitors are potential cancer therapeutics that can be utilized for enhancing the efficacy of DNA damaging agents. Multiple small molecule CHK1 inhibitors from different chemical scaffolds have been developed and evaluated in clinical trials in combination with chemotherapeutics and radiation treatment. Scaffold morphing of thiophene carboxamide ureas (TCUs), such as AZD7762 (1) and a related series of triazoloquinolines (TZQs), led to the identification of fused-ring bicyclic CHK1 inhibitors, 7-carboxamide thienopyridines (7-CTPs), and 7-carboxamide indoles. X-ray crystal structures reveal a key intramolecular noncovalent sulfur-oxygen interaction in aligning the hinge-binding carboxamide group to the thienopyridine core in a coplanar fashion. An intramolecular hydrogen bond to an indole NH was also effective in locking the carboxamide in the preferred bound conformation to CHK1. Optimization on the 7-CTP series resulted in the identification of lead compound 44, which displayed respectable drug-like properties and good in vitro and in vivo potency.

  11. EMODnet MedSea Checkpoint for sustainable Blue Growth

    Science.gov (United States)

    Moussat, Eric; Pinardi, Nadia; Manzella, Giuseppe; Blanc, Frederique

    2016-04-01

    The EMODNET checkpoint is a wide monitoring system assessment activity aiming to support the sustainable Blue Growth at the scale of the European Sea Basins by: 1) Clarifying the observation landscape of all compartments of the marine environment including Air, Water, Seabed, Biota and Human activities, pointing out to the existing programs, national, European and international 2) Evaluating fitness for use indicators that will show the accessibility and usability of observation and modeling data sets and their roles and synergies based upon selected applications by the European Marine Environment Strategy 3) Prioritizing the needs to optimize the overall monitoring Infrastructure (in situ and satellite data collection and assembling, data management and networking, modeling and forecasting, geo-infrastructure) and release recommendations for evolutions to better meet the application requirements in view of sustainable Blue Growth The assessment is designed for : - Institutional stakeholders for decision making on observation and monitoring systems - Data providers and producers to know how their data collected once for a given purpose could fit other user needs - End-users interested in a regional status and possible uses of existing monitoring data Selected end-user applications are of paramount importance for: (i) the blue economy sector (offshore industries, fisheries); (ii) marine environment variability and change (eutrophication, river inputs and ocean climate change impacts); (iii) emergency management (oil spills); and (iv) preservation of natural resources and biodiversity (Marine Protected Areas). End-user applications generate innovative products based on the existing observation landscape. The fitness for use assessment is made thanks to the comparison of the expected product specifications with the quality of the product derived from the selected data. This involves the development of checkpoint information and indicators based on Data quality and

  12. Acute inflammatory thyromegaly following checkpoint inhibition: A new imaging entity?

    Directory of Open Access Journals (Sweden)

    Erik H. Middlebrooks, MD

    2018-02-01

    Full Text Available Immune checkpoint blockade (CPB utilizing such agents as ipilimumab, nivolumab, or pembrolizumab has revolutionized melanoma therapy and has seen continued utilization in numerous other malignancies in recent years. However, these agents come at the price of inflammatory immune-related adverse events. Despite the increasing recognition of biochemical thyroid dysfunction associated with CPB, information regarding potential imaging findings is sparse. We describe the first 2 cases of acute thyroiditis following CPB presenting as diffuse thyromegaly documented by computed tomography, ultrasound, and iodine uptake imaging. Given the rise in the use of CPB, it is important for radiologists to recognize potential imaging manifestations of therapy immune-related adverse events to avoid erroneous diagnosis and to prompt the biochemical investigation of thyroid function.

  13. Role of Immune Checkpoint Inhibitors in Small Cell Lung Cancer.

    Science.gov (United States)

    Cooper, Maryann R; Alrajhi, Abdullah M; Durand, Cheryl R

    Small cell lung cancer (SCLC) accounts for approximately 13% of all lung cancer diagnoses each year. SCLC is characterized by a rapid doubling time, early metastatic spread, and an unfavorable prognosis overall. Most patients with SCLC will respond to initial treatment; however, the majority will experience a disease recurrence and response to second-line therapies is poor. Immune checkpoint inhibitors may be an option given the success in other diseases. A literature search was conducted using Medline (1946-July week 1, 2017) and Embase (1996-2017 week 28) with the search terms small cell lung cancer combined with nivolumab or ipilimumab or pembrolizumab or atezolizumab or tremelimumab or durvalumab. Five clinical trials, including extended follow-up for 2, that evaluated immune checkpoint inhibitors in limited stage or extensive stage SCLC were included. In 2 phase 2 trials, ipilimumab was added to upfront chemotherapy. In both trials, an improvement in progression-free survival was seen. Toxicity, when combined with a platinum and etoposide, was significant. In a confirmatory phase 3 trial, ipilimumab did not prolong overall survival when added to first-line chemotherapy. Overall, response rates were similar between the placebo and ipilimumab groups. A phase 1/2 trial evaluated nivolumab alone or in combination with ipilimumab in recurrent SCLC. Results revealed that nivolumab monotherapy and the combination of nivolumab and ipilimumab were relatively safe and had antitumor activity. Pembrolizumab has been evaluated in a multicohort, phase 1b trial. Preliminary data showed a durable response in the second-line setting. Given the lack of overall survival data and significant toxicity associated with the combination of ipilimumab with first-line chemotherapy, this treatment is not a reasonable option at this time. Nivolumab alone or in combination with ipilimumab is a valid option for recurrent SCLC.

  14. Tousled-like kinase-dependent phosphorylation of Rad9 plays a role in cell cycle progression and G2/M checkpoint exit.

    Directory of Open Access Journals (Sweden)

    Ryan Kelly

    Full Text Available Genomic integrity is preserved by checkpoints, which act to delay cell cycle progression in the presence of DNA damage or replication stress. The heterotrimeric Rad9-Rad1-Hus1 (9-1-1 complex is a PCNA-like clamp that is loaded onto DNA at structures resulting from damage and is important for initiating and maintaining the checkpoint response. Rad9 possesses a C-terminal tail that is phosphorylated constitutively and in response to cell cycle position and DNA damage. Previous studies have identified tousled-like kinase 1 (TLK1 as a kinase that may modify Rad9. Here we show that Rad9 is phosphorylated in a TLK-dependent manner in vitro and in vivo, and that T355 within the C-terminal tail is the primary targeted residue. Phosphorylation of Rad9 at T355 is quickly reduced upon exposure to ionizing radiation before returning to baseline later in the damage response. We also show that TLK1 and Rad9 interact constitutively, and that this interaction is enhanced in chromatin-bound Rad9 at later stages of the damage response. Furthermore, we demonstrate via siRNA-mediated depletion that TLK1 is required for progression through S-phase in normally cycling cells, and that cells lacking TLK1 display a prolonged G2/M arrest upon exposure to ionizing radiation, a phenotype that is mimicked by over-expression of a Rad9-T355A mutant. Given that TLK1 has previously been shown to be transiently inactivated upon phosphorylation by Chk1 in response to DNA damage, we propose that TLK1 and Chk1 act in concert to modulate the phosphorylation status of Rad9, which in turn serves to regulate the DNA damage response.

  15. The pachytene checkpoint and its relationship to evolutionary patterns of polyploidization and hybrid sterility.

    Science.gov (United States)

    Li, X C; Barringer, B C; Barbash, D A

    2009-01-01

    Sterility is a commonly observed phenotype in interspecific hybrids. Sterility may result from chromosomal or genic incompatibilities, and much progress has been made toward understanding the genetic basis of hybrid sterility in various taxa. The underlying mechanisms causing hybrid sterility, however, are less well known. The pachytene checkpoint is a meiotic surveillance system that many organisms use to detect aberrant meiotic products, in order to prevent the production of defective gametes. We suggest that activation of the pachytene checkpoint may be an important mechanism contributing to two types of hybrid sterility. First, the pachytene checkpoint may form the mechanistic basis of some gene-based hybrid sterility phenotypes. Second, the pachytene checkpoint may be an important mechanism that mediates chromosomal-based hybrid sterility phenotypes involving gametes with non-haploid (either non-reduced or aneuploid) chromosome sets. Studies in several species suggest that the strength of the pachytene checkpoint is sexually dimorphic, observations that warrant future investigation into whether such variation may contribute to differences in patterns of sterility between male and female interspecific hybrids. In addition, plants seem to lack the pachytene checkpoint, which correlates with increased production of unreduced gametes and a higher incidence of polyploid species in plants versus animals. Although the pachytene checkpoint occurs in many animals and in fungi, at least some of the genes that execute the pachytene checkpoint are different among organisms. This finding suggests that the penetrance of the pachytene checkpoint, and even its presence or absence can evolve rapidly. The surprising degree of evolutionary flexibility in this meiotic surveillance system may contribute to the observed variation in patterns of hybrid sterility and in rates of polyploidization.

  16. The DNA damage checkpoint precedes activation of ARF in response to escalating oncogenic stress during tumorigenesis

    DEFF Research Database (Denmark)

    Evangelou, K.; Bartkova, J.; Kotsinas, A.

    2013-01-01

    oncogenes showed that the delayed upregulation of ARF reflected a requirement for a higher, transcriptionally based threshold of oncogenic stress, elicited by at least two oncogenic 'hits', compared with lower activation threshold for DDR. We propose that relative to DDR activation, ARF provides...

  17. The DNA damage checkpoint precedes activation of ARF in response to escalating oncogenic stress during tumorigenesis

    Czech Academy of Sciences Publication Activity Database

    Evangelou, K.; Bartkova, J.; Kotsinas, A.; Pateras, I.S.; Liontos, M.; Velimezi, G.; Košař, Martin; Liloglou, T.; Trougakos, I.P.; Dyrskjot, L.; Andersen, C.J.; Papaioannou, G.; Drosos, A.; Papafotiou, M.; Hodný, Zdeněk; Sosa-Pineda, B.; Wu, X.R.; Klinakis, A.; Orntoft, T.; Lukas, J.; Bartek, Jiří; Gorgoulis, V.G.

    2013-01-01

    Roč. 20, č. 11 (2013), s. 1485-1497 ISSN 1350-9047 Institutional support: RVO:68378050 Keywords : ARF * carcinogenesis * DDR * E2F1 * p16(INK4A) Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.385, year: 2013

  18. Polo-like kinase-1 is a target of the DNA damage checkpoint

    NARCIS (Netherlands)

    Smits, V.A.J.; Klompmaker, R.; Arnaud, L.; Rijksen, G.; Nigg, E.A.; Medema, R.H.

    2000-01-01

    Polo-like kinases (PLKs) have an important role in several stages of mitosis. They contribute to the activation of cyclin B/Cdc2 and are involved in centrosome maturation and bipolar spindle formation at the onset of mitosis1, 2. PLKs also control mitotic exit by regulating the anaphase-promoting

  19. An ATM-independent S-phase checkpoint response involves CHK1 pathway

    Science.gov (United States)

    Zhou, Xiang-Yang; Wang, Xiang; Hu, Baocheng; Guan, Jun; Iliakis, George; Wang, Ya

    2002-01-01

    After exposure to genotoxic stress, proliferating cells actively slow down the DNA replication through a S-phase checkpoint to provide time for repair. We report that in addition to the ataxia-telangiectasia mutated (ATM)-dependent pathway that controls the fast response, there is an ATM-independent pathway that controls the slow response to regulate the S-phase checkpoint after ionizing radiation in mammalian cells. The slow response of S-phase checkpoint, which is resistant to wortmannin, sensitive to caffeine and UCN-01, and related to cyclin-dependent kinase phosphorylation, is much stronger in CHK1 overexpressed cells, and it could be abolished by Chk1 antisense oligonucleotides. These results provide evidence that the ATM-independent slow response of S-phase checkpoint involves CHK1 pathway.

  20. Immune-checkpoint inhibitors in the era of precision medicine: What radiologists should know

    Energy Technology Data Exchange (ETDEWEB)

    Braschi-Amirfarzan, Marta; Tirumani, Sree Harsha; Hodi, Frank Stephan Jr; Nishno, Mizuki [Dept. of Radiology, Brigham and Women' s Hospital and Dana Farber Cancer Institute, Boston (United States)

    2017-01-15

    Over the past five years immune-checkpoint inhibitors have dramatically changed the therapeutic landscape of advanced solid and hematologic malignancies. The currently approved immune-checkpoint inhibitors include antibodies to cytotoxic T-lymphocyte antigen-4, programmed cell death (PD-1), and programmed cell death ligand (PD-L1 and PD-L2). Response to immune-checkpoint inhibitors is evaluated on imaging using the immune-related response criteria. Activation of immune system results in a unique toxicity profile termed immune-related adverse events. This article will review the molecular mechanism, clinical applications, imaging of immune-related response patterns and adverse events associated with immune-checkpoint inhibitors.

  1. Immune-checkpoint inhibitors in the era of precision medicine: What radiologists should know

    International Nuclear Information System (INIS)

    Braschi-Amirfarzan, Marta; Tirumani, Sree Harsha; Hodi, Frank Stephan Jr; Nishno, Mizuki

    2017-01-01

    Over the past five years immune-checkpoint inhibitors have dramatically changed the therapeutic landscape of advanced solid and hematologic malignancies. The currently approved immune-checkpoint inhibitors include antibodies to cytotoxic T-lymphocyte antigen-4, programmed cell death (PD-1), and programmed cell death ligand (PD-L1 and PD-L2). Response to immune-checkpoint inhibitors is evaluated on imaging using the immune-related response criteria. Activation of immune system results in a unique toxicity profile termed immune-related adverse events. This article will review the molecular mechanism, clinical applications, imaging of immune-related response patterns and adverse events associated with immune-checkpoint inhibitors

  2. Dihydrocoumarin, an HDAC Inhibitor, Increases DNA Damage Sensitivity by Inhibiting Rad52

    Directory of Open Access Journals (Sweden)

    Chin-Chuan Chen

    2017-12-01

    Full Text Available Effective DNA repair enables cancer cells to survive DNA damage induced by chemotherapeutic or radiotherapeutic treatments. Therefore, inhibiting DNA repair pathways is a promising therapeutic strategy for increasing the efficacy of such treatments. In this study, we found that dihydrocoumarin (DHC, a flavoring agent, causes deficiencies in double-stand break (DSB repair and prolonged DNA damage checkpoint recovery in yeast. Following DNA damage, Rad52 recombinase was revealed to be inhibited by DHC, which results in deficiencies in DSB repair and prolonged DNA damage checkpoint recovery. The deletion of RPD3, a class I histone deacetylase (HDAC, was found to mimic DHC-induced suppression of Rad52 expression, suggesting that the HDAC inhibitor activity of DHC is critical to DSB repair and DNA damage sensitivity. Overall, our findings delineate the regulatory mechanisms of DHC in DSB repair and suggest that it might potentially be used as an inhibitor of the DNA repair pathway in human cells.

  3. Resistance to checkpoint blockade therapy through inactivation of antigen presentation.

    Science.gov (United States)

    Sade-Feldman, Moshe; Jiao, Yunxin J; Chen, Jonathan H; Rooney, Michael S; Barzily-Rokni, Michal; Eliane, Jean-Pierre; Bjorgaard, Stacey L; Hammond, Marc R; Vitzthum, Hans; Blackmon, Shauna M; Frederick, Dennie T; Hazar-Rethinam, Mehlika; Nadres, Brandon A; Van Seventer, Emily E; Shukla, Sachet A; Yizhak, Keren; Ray, John P; Rosebrock, Daniel; Livitz, Dimitri; Adalsteinsson, Viktor; Getz, Gad; Duncan, Lyn M; Li, Bo; Corcoran, Ryan B; Lawrence, Donald P; Stemmer-Rachamimov, Anat; Boland, Genevieve M; Landau, Dan A; Flaherty, Keith T; Sullivan, Ryan J; Hacohen, Nir

    2017-10-26

    Treatment with immune checkpoint blockade (CPB) therapies often leads to prolonged responses in patients with metastatic melanoma, but the common mechanisms of primary and acquired resistance to these agents remain incompletely characterized and have yet to be validated in large cohorts. By analyzing longitudinal tumor biopsies from 17 metastatic melanoma patients treated with CPB therapies, we observed point mutations, deletions or loss of heterozygosity (LOH) in beta-2-microglobulin (B2M), an essential component of MHC class I antigen presentation, in 29.4% of patients with progressing disease. In two independent cohorts of melanoma patients treated with anti-CTLA4 and anti-PD1, respectively, we find that B2M LOH is enriched threefold in non-responders (~30%) compared to responders (~10%) and associated with poorer overall survival. Loss of both copies of B2M is found only in non-responders. B2M loss is likely a common mechanism of resistance to therapies targeting CTLA4 or PD1.

  4. Identification of a novel EGF-sensitive cell cycle checkpoint

    International Nuclear Information System (INIS)

    Walker, Francesca; Zhang Huihua; Burgess, Antony W.

    2007-01-01

    The site of action of growth factors on mammalian cell cycle has been assigned to the boundary between the G1 and S phases. We show here that Epidermal Growth Factor (EGF) is also required for mitosis. BaF/3 cells expressing the EGFR (BaF/wtEGFR) synthesize DNA in response to EGF, but arrest in S-phase. We have generated a cell line (BaF/ERX) with defective downregulation of the EGFR and sustained activation of EGFR signalling pathways: these cells undergo mitosis in an EGF-dependent manner. The transit of BaF/ERX cells through G2/M strictly requires activation of EGFR and is abolished by AG1478. This phenotype is mimicked by co-expression of ErbB2 in BaF/wtEGFR cells, and abolished by inhibition of the EGFR kinase, suggesting that sustained signalling of the EGFR, through impaired downregulation of the EGFR or heterodimerization, is required for completion of the cycle. We have confirmed the role of EGFR signalling in the G2/M phase of the cell cycle using a human tumor cell line which overexpresses the EGFR and is dependent on EGFR signalling for growth. These findings unmask an EGF-sensitive checkpoint, helping to understand the link between sustained EGFR signalling, proliferation and the acquisition of a radioresistant phenotype in cancer cells

  5. Loss of the Greatwall Kinase Weakens the Spindle Assembly Checkpoint.

    Directory of Open Access Journals (Sweden)

    M Kasim Diril

    2016-09-01

    Full Text Available The Greatwall kinase/Mastl is an essential gene that indirectly inhibits the phosphatase activity toward mitotic Cdk1 substrates. Here we show that although Mastl knockout (MastlNULL MEFs enter mitosis, they progress through mitosis without completing cytokinesis despite the presence of misaligned chromosomes, which causes chromosome segregation defects. Furthermore, we uncover the requirement of Mastl for robust spindle assembly checkpoint (SAC maintenance since the duration of mitotic arrest caused by microtubule poisons in MastlNULL MEFs is shortened, which correlates with premature disappearance of the essential SAC protein Mad1 at the kinetochores. Notably, MastlNULL MEFs display reduced phosphorylation of a number of proteins in mitosis, which include the essential SAC kinase MPS1. We further demonstrate that Mastl is required for multi-site phosphorylation of MPS1 as well as robust MPS1 kinase activity in mitosis. In contrast, treatment of MastlNULL cells with the phosphatase inhibitor okadaic acid (OKA rescues the defects in MPS1 kinase activity, mislocalization of phospho-MPS1 as well as Mad1 at the kinetochore, and premature SAC silencing. Moreover, using in vitro dephosphorylation assays, we demonstrate that Mastl promotes persistent MPS1 phosphorylation by inhibiting PP2A/B55-mediated MPS1 dephosphorylation rather than affecting Cdk1 kinase activity. Our findings establish a key regulatory function of the Greatwall kinase/Mastl->PP2A/B55 pathway in preventing premature SAC silencing.

  6. Loss of the Greatwall Kinase Weakens the Spindle Assembly Checkpoint.

    Science.gov (United States)

    Diril, M Kasim; Bisteau, Xavier; Kitagawa, Mayumi; Caldez, Matias J; Wee, Sheena; Gunaratne, Jayantha; Lee, Sang Hyun; Kaldis, Philipp

    2016-09-01

    The Greatwall kinase/Mastl is an essential gene that indirectly inhibits the phosphatase activity toward mitotic Cdk1 substrates. Here we show that although Mastl knockout (MastlNULL) MEFs enter mitosis, they progress through mitosis without completing cytokinesis despite the presence of misaligned chromosomes, which causes chromosome segregation defects. Furthermore, we uncover the requirement of Mastl for robust spindle assembly checkpoint (SAC) maintenance since the duration of mitotic arrest caused by microtubule poisons in MastlNULL MEFs is shortened, which correlates with premature disappearance of the essential SAC protein Mad1 at the kinetochores. Notably, MastlNULL MEFs display reduced phosphorylation of a number of proteins in mitosis, which include the essential SAC kinase MPS1. We further demonstrate that Mastl is required for multi-site phosphorylation of MPS1 as well as robust MPS1 kinase activity in mitosis. In contrast, treatment of MastlNULL cells with the phosphatase inhibitor okadaic acid (OKA) rescues the defects in MPS1 kinase activity, mislocalization of phospho-MPS1 as well as Mad1 at the kinetochore, and premature SAC silencing. Moreover, using in vitro dephosphorylation assays, we demonstrate that Mastl promotes persistent MPS1 phosphorylation by inhibiting PP2A/B55-mediated MPS1 dephosphorylation rather than affecting Cdk1 kinase activity. Our findings establish a key regulatory function of the Greatwall kinase/Mastl->PP2A/B55 pathway in preventing premature SAC silencing.

  7. DNA Checkpoint and Repair Factors Are Nuclear Sensors for Intracellular Organelle Stresses—Inflammations and Cancers Can Have High Genomic Risks

    Directory of Open Access Journals (Sweden)

    Huihong Zeng

    2018-05-01

    Full Text Available Under inflammatory conditions, inflammatory cells release reactive oxygen species (ROS and reactive nitrogen species (RNS which cause DNA damage. If not appropriately repaired, DNA damage leads to gene mutations and genomic instability. DNA damage checkpoint factors (DDCF and DNA damage repair factors (DDRF play a vital role in maintaining genomic integrity. However, how DDCFs and DDRFs are modulated under physiological and pathological conditions are not fully known. We took an experimental database analysis to determine the expression of 26 DNA DDCFs and 42 DNA DDRFs in 21 human and 20 mouse tissues in physiological/pathological conditions. We made the following significant findings: (1 Few DDCFs and DDRFs are ubiquitously expressed in tissues while many are differentially regulated.; (2 the expression of DDCFs and DDRFs are modulated not only in cancers but also in sterile inflammatory disorders and metabolic diseases; (3 tissue methylation status, pro-inflammatory cytokines, hypoxia regulating factors and tissue angiogenic potential can determine the expression of DDCFs and DDRFs; (4 intracellular organelles can transmit the stress signals to the nucleus, which may modulate the cell death by regulating the DDCF and DDRF expression. Our results shows that sterile inflammatory disorders and cancers increase genomic instability, therefore can be classified as pathologies with a high genomic risk. We also propose a new concept that as parts of cellular sensor cross-talking network, DNA checkpoint and repair factors serve as nuclear sensors for intracellular organelle stresses. Further, this work would lead to identification of novel therapeutic targets and new biomarkers for diagnosis and prognosis of metabolic diseases, inflammation, tissue damage and cancers.

  8. Immune checkpoint inhibitor-induced gastrointestinal and hepatic injury: pathologists' perspective.

    Science.gov (United States)

    Karamchandani, Dipti M; Chetty, Runjan

    2018-04-27

    Immune checkpoint inhibitors (CPIs) are a relatively new class of 'miracle' dugs that have revolutionised the treatment and prognosis of some advanced-stage malignancies, and have increased the survival rates significantly. This class of drugs includes cytotoxic T lymphocyte antigen-4 inhibitors such as ipilimumab; programmed cell death protein-1 inhibitors such as nivolumab, pembrolizumab and avelumab; and programmed cell death protein ligand-1 inhibitors such as atezolizumab. These drugs stimulate the immune system by blocking the coinhibitory receptors on the T cells and lead to antitumoural response. However, a flip side of these novel drugs is immune-related adverse events (irAEs), secondary to immune-mediated process due to disrupted self-tolerance. The irAEs in the gastrointestinal (GI) tract/liver may result in diarrhoea, colitis or hepatitis. An accurate diagnosis of CPI-induced colitis and/or hepatitis is essential for optimal patient management. As we anticipate greater use of these drugs in the future given the significant clinical response, pathologists need to be aware of the spectrum of histological findings that may be encountered in GI and/or liver biopsies received from these patients, as well as differentiate them from its histopathological mimics. This present review discusses the clinical features, detailed histopathological features, management and the differential diagnosis of the luminal GI and hepatic irAEs that may be encountered secondary to CPI therapy. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  9. DW-MRI as a Predictive Biomarker of Radiosensitization of GBM through Targeted Inhibition of Checkpoint Kinases.

    Science.gov (United States)

    Williams, Terence M; Galbán, Stefanie; Li, Fei; Heist, Kevin A; Galbán, Craig J; Lawrence, Theodore S; Holland, Eric C; Thomae, Tami L; Chenevert, Thomas L; Rehemtulla, Alnawaz; Ross, Brian D

    2013-04-01

    The inherent treatment resistance of glioblastoma (GBM) can involve multiple mechanisms including checkpoint kinase (Chk1/2)-mediated increased DNA repair capability, which can attenuate the effects of genotoxic chemotherapies and radiation. The goal of this study was to evaluate diffusion-weighted magnetic resonance imaging (DW-MRI) as a biomarker for Chk1/2 inhibitors in combination with radiation for enhancement of treatment efficacy in GBM. We evaluated a specific small molecule inhibitor of Chk1/2, AZD7762, in combination with radiation using in vitro human cell lines and in vivo using a genetically engineered GBM mouse model. DW-MRI and T1-contrast MRI were used to follow treatment effects on intracranial tumor cellularity and growth rates, respectively. AZD7762 inhibited clonal proliferation in a panel of GBM cell lines and increased radiosensitivity in p53-mutated GBM cell lines to a greater extent compared to p53 wild-type cells. In vivo efficacy of AZD7762 demonstrated a dose-dependent inhibitory effect on GBM tumor growth rate and a reduction in tumor cellularity based on DW-MRI scans along with enhancement of radiation efficacy. DW-MRI was found to be a useful imaging biomarker for the detection of radiosensitization through inhibition of checkpoint kinases. Chk1/2 inhibition resulted in antiproliferative activity, prevention of DNA damage-induced repair, and radiosensitization in preclinical GBM tumor models, both in vitro and in vivo. The effects were found to be maximal in p53-mutated GBM cells. These results provide the rationale for integration of DW-MRI in clinical translation of Chk1/2 inhibition with radiation for the treatment of GBM.

  10. Residual Cdk1/2 activity after DNA damage promotes senescence

    Czech Academy of Sciences Publication Activity Database

    Müllers, E.; Cascales, H.S.; Burdová, Kamila; Macůrek, Libor; Lindqvist, A.

    2017-01-01

    Roč. 16, č. 3 (2017), s. 575-584 ISSN 1474-9726 R&D Projects: GA ČR GA13-18392S Institutional support: RVO:68378050 Keywords : Cdk1 * Cdk2 * cell cycle * checkpoint recovery * DNA damage response * G2phase * p21 * senescence Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biochemistry and molecular biology

  11. Adrenergic Signaling: A Targetable Checkpoint Limiting Development of the Antitumor Immune Response

    Science.gov (United States)

    Qiao, Guanxi; Chen, Minhui; Bucsek, Mark J.; Repasky, Elizabeth A.; Hylander, Bonnie L.

    2018-01-01

    An immune response must be tightly controlled so that it will be commensurate with the level of response needed to protect the organism without damaging normal tissue. The roles of cytokines and chemokines in orchestrating these processes are well known, but although stress has long been thought to also affect immune responses, the underlying mechanisms were not as well understood. Recently, the role of nerves and, specifically, the sympathetic nervous system, in regulating immune responses is being revealed. Generally, an acute stress response is beneficial but chronic stress is detrimental because it suppresses the activities of effector immune cells while increasing the activities of immunosuppressive cells. In this review, we first discuss the underlying biology of adrenergic signaling in cells of both the innate and adaptive immune system. We then focus on the effects of chronic adrenergic stress in promoting tumor growth, giving examples of effects on tumor cells and immune cells, explaining the methods commonly used to induce stress in preclinical mouse models. We highlight how this relates to our observations that mandated housing conditions impose baseline chronic stress on mouse models, which is sufficient to cause chronic immunosuppression. This problem is not commonly recognized, but it has been shown to impact conclusions of several studies of mouse physiology and mouse models of disease. Moreover, the fact that preclinical mouse models are chronically immunosuppressed has critical ramifications for analysis of any experiments with an immune component. Our group has found that reducing adrenergic stress by housing mice at thermoneutrality or treating mice housed at cooler temperatures with β-blockers reverses immunosuppression and significantly improves responses to checkpoint inhibitor immunotherapy. These observations are clinically relevant because there are numerous retrospective epidemiological studies concluding that cancer patients who were

  12. Smurf2 as a novel mitotic regulator: From the spindle assembly checkpoint to tumorigenesis

    Directory of Open Access Journals (Sweden)

    Moore Finola E

    2009-07-01

    Full Text Available Abstract The execution of the mitotic program with high fidelity is dependent upon precise spatiotemporal regulation of posttranslational protein modifications. For example, the timely polyubiquitination of critical mitotic regulators by Anaphase Promoting Complex/Cyclosome (APC/C is essential for the metaphase to anaphase transition and mitotic exit. The spindle assembly checkpoint prevents unscheduled activity of APC/C-Cdc20 in early mitosis, allowing bipolar attachment of kinetochores to mitotic spindle and facilitating equal segregation of sister chromatids. The critical effector of the spindle checkpoint, Mitotic arrest deficient 2 (Mad2, is recruited to unattached kinetochores forming a complex with other regulatory proteins to efficiently and cooperatively inhibit APC/C-Cdc20. A weakened and/or dysfunctional spindle checkpoint has been linked to the development of genomic instability in both cell culture and animal models, and evidence suggests that aberrant regulation of the spindle checkpoint plays a critical role in human carcinogenesis. Recent studies have illuminated a network of both degradative and non-degradative ubiquitination events that regulate the metaphase to anaphase transition and mitotic exit. Within this context, our recent work showed that the HECT (Homologous to E6-AP C-terminus-family E3 ligase Smurf2 (Smad specific ubiquitin regulatory factor 2, known as a negative regulator of transforming growth factor-beta (TGF-β signaling, is required for a functional spindle checkpoint by promoting the functional localization and stability of Mad2. Here we discuss putative models explaining the role of Smurf2 as a new regulator in the spindle checkpoint. The dynamic mitotic localization of Smurf2 to the centrosome and other critical mitotic structures provides implications about mitotic checkpoint control dependent on various ubiquitination events. Finally, deregulated Smurf2 activity may contribute to carcinogenesis by

  13. Asynchronous Two-Level Checkpointing Scheme for Large-Scale Adjoints in the Spectral-Element Solver Nek5000

    Energy Technology Data Exchange (ETDEWEB)

    Schanen, Michel; Marin, Oana; Zhang, Hong; Anitescu, Mihai

    2016-01-01

    Adjoints are an important computational tool for large-scale sensitivity evaluation, uncertainty quantification, and derivative-based optimization. An essential component of their performance is the storage/recomputation balance in which efficient checkpointing methods play a key role. We introduce a novel asynchronous two-level adjoint checkpointing scheme for multistep numerical time discretizations targeted at large-scale numerical simulations. The checkpointing scheme combines bandwidth-limited disk checkpointing and binomial memory checkpointing. Based on assumptions about the target petascale systems, which we later demonstrate to be realistic on the IBM Blue Gene/Q system Mira, we create a model of the expected performance of our checkpointing approach and validate it using the highly scalable Navier-Stokes spectralelement solver Nek5000 on small to moderate subsystems of the Mira supercomputer. In turn, this allows us to predict optimal algorithmic choices when using all of Mira. We also demonstrate that two-level checkpointing is significantly superior to single-level checkpointing when adjoining a large number of time integration steps. To our knowledge, this is the first time two-level checkpointing had been designed, implemented, tuned, and demonstrated on fluid dynamics codes at large scale of 50k+ cores.

  14. Casein kinase II is required for the spindle assembly checkpoint by regulating Mad2p in fission yeast

    International Nuclear Information System (INIS)

    Shimada, Midori; Yamamoto, Ayumu; Murakami-Tonami, Yuko; Nakanishi, Makoto; Yoshida, Takashi; Aiba, Hirofumi; Murakami, Hiroshi

    2009-01-01

    The spindle checkpoint is a surveillance mechanism that ensures the fidelity of chromosome segregation in mitosis. Here we show that fission yeast casein kinase II (CK2) is required for this checkpoint function. In the CK2 mutants mitosis occurs in the presence of a spindle defect, and the spindle checkpoint protein Mad2p fails to localize to unattached kinetochores. The CK2 mutants are sensitive to the microtubule depolymerising drug thiabendazole, which is counteracted by ectopic expression of mad2 + . The level of Mad2p is low in the CK2 mutants. These results suggest that CK2 has a role in the spindle checkpoint by regulating Mad2p.

  15. In-silico modeling of the mitotic spindle assembly checkpoint.

    Directory of Open Access Journals (Sweden)

    Bashar Ibrahim

    2008-02-01

    Full Text Available The Mitotic Spindle Assembly Checkpoint ((MSAC is an evolutionary conserved mechanism that ensures the correct segregation of chromosomes by restraining cell cycle progression from entering anaphase until all chromosomes have made proper bipolar attachments to the mitotic spindle. Its malfunction can lead to cancer.We have constructed and validated for the human (MSAC mechanism an in silico dynamical model, integrating 11 proteins and complexes. The model incorporates the perspectives of three central control pathways, namely Mad1/Mad2 induced Cdc20 sequestering based on the Template Model, MCC formation, and APC inhibition. Originating from the biochemical reactions for the underlying molecular processes, non-linear ordinary differential equations for the concentrations of 11 proteins and complexes of the (MSAC are derived. Most of the kinetic constants are taken from literature, the remaining four unknown parameters are derived by an evolutionary optimization procedure for an objective function describing the dynamics of the APC:Cdc20 complex. MCC:APC dissociation is described by two alternatives, namely the "Dissociation" and the "Convey" model variants. The attachment of the kinetochore to microtubuli is simulated by a switching parameter silencing those reactions which are stopped by the attachment. For both, the Dissociation and the Convey variants, we compare two different scenarios concerning the microtubule attachment dependent control of the dissociation reaction. Our model is validated by simulation of ten perturbation experiments.Only in the controlled case, our models show (MSAC behaviour at meta- to anaphase transition in agreement with experimental observations. Our simulations revealed that for (MSAC activation, Cdc20 is not fully sequestered; instead APC is inhibited by MCC binding.

  16. Acute symptomatic hypocalcemia from immune checkpoint therapy-induced hypoparathyroidism.

    Science.gov (United States)

    Win, Myint Aung; Thein, Kyaw Zin; Qdaisat, Aiham; Yeung, Sai-Ching Jim

    2017-07-01

    Ipilimumab (a monoclonal antibody against CTLA-4) and nivolumab (a humanized antibody against PD-1) target these immune checkpoint pathways and are used for treatment of melanoma and an increasing number of other cancers. However, they may cause immune-related adverse effects (IRAEs). Although many endocrinopathies are known to be IRAEs, primary hypoparathyroidism with severe hypocalcemia has never been reported. This is the first case of hypoparathyroidism as an IRAE presenting to an Emergency Department with acute hypocalcemia. A 73-year-old man with metastatic melanoma presented to the Emergency Department for the chief complaints of imbalance, general muscle weakness, abdominal pain and tingling in extremities. He had wide spread metastasis, and begun immunotherapy with concurrent ipilimumab and nivolumab 1.5months ago. At presentation, he had ataxia, paresthesia in the hands and feet, and abdominal cramping. Magnetic resonance imaging of the brain was unremarkable. He was found to be hypocalcemic with undetectable plasma parathyroid hormone. He was admitted for treatment of symptomatic hypocalcemia and was diagnosed with primary hypoparathyroidism. Shortly afterwards, he had thyrotoxicosis manifesting as tachycardia and anxiety, followed by development of primary hypothyroidism. At 4months after the Emergency Department visit, his parathyroid function and thyroid function had not recovered, and required continued thyroid hormone replacement and calcium and vitamin D treatment for hypocalcemia. Primary hypoparathyroidism caused by ipilimumab and nivolumab may acute manifest with severe symptomatic hypocalcemia. Emergency care providers should be aware of hypoparathyroidism as a new IRAE in this new era of immuno-oncology. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Calreticulin reveals a critical Ca2+ checkpoint in cardiac myofibrillogenesis

    Science.gov (United States)

    Li, Jian; Pucéat, Michel; Perez-Terzic, Carmen; Mery, Annabelle; Nakamura, Kimitoshi; Michalak, Marek; Krause, Karl-Heinz; Jaconi, Marisa E.

    2002-01-01

    Calreticulin (crt) is an ubiquitously expressed and multifunctional Ca2+-binding protein that regulates diverse vital cell functions, including Ca2+ storage in the ER and protein folding. Calreticulin deficiency in mice is lethal in utero due to defects in heart development and function. Herein, we used crt − / − embryonic stem (ES) cells differentiated in vitro into cardiac cells to investigate the molecular mechanisms underlying heart failure of knockout embryos. After 8 d of differentiation, beating areas were prominent in ES-derived wild-type (wt) embryoid bodies (EBs), but not in ES-derived crt − / − EBs, despite normal expression levels of cardiac transcription factors. Crt − / − EBs exhibited a severe decrease in expression and a lack of phosphorylation of ventricular myosin light chain 2 (MLC2v), resulting in an impaired organization of myofibrils. Crt − / − phenotype could be recreated in wt cells by chelating extracellular or cytoplasmic Ca2+ with EGTA or BAPTA, or by inhibiting Ca2+/calmodulin-dependent kinases (CaMKs). An imposed ionomycin-triggered cystolic-free Ca2+ concentration ([Ca2+]c) elevation restored the expression, phosphorylation, and insertion of MLC2v into sarcomeric structures and in turn the myofibrillogenesis. The transcription factor myocyte enhancer factor C2 failed to accumulate into nuclei of crt − / − cardiac cells in the absence of ionomycin-triggered [Ca2+]c increase. We conclude that the absence of calreticulin interferes with myofibril formation. Most importantly, calreticulin deficiency revealed the importance of a Ca2+-dependent checkpoint critical for early events during cardiac myofibrillogenesis. PMID:12105184

  18. Checkpoint inhibitors in advanced melanoma: effect on the field of immunotherapy.

    Science.gov (United States)

    O'reilly, Aine; Larkin, James

    2017-07-01

    The success of the immune checkpoint inhibitors in melanoma has reinvigorated the field of immunotherapy. Immune checkpoint inhibitors are now the standard of care in multiple cancer types including lung cancer, head and neck cancer, urothelial cancer and renal cell cancer. The field of immunotherapy is currently expanding rapidly and will be a focus of research and development for decades to come. Areas covered: This review covers the early development of immune checkpoint inhibitors and the changes that occurred in the drug development paradigm to facilitate the development of immunotherapy. The review will summarise the areas into which immune checkpoint inhibitors have been adopted and will review the data that supported this. Furthermore, we will discuss future developments in immunotherapy and the current landscape regarding maximising the potential of immunotherapy in clinical practice. Expert commentary: In the author's opinion, the potential of immunotherapy is vast. To date immune checkpoint inhibition has already delivered durable responses in a proportion of patients with cancer types which were previously universally lethal. The future of immunotherapy will rely upon the intelligent application of translational research to clinical practice, such that immunotherapy can be effective for a wider population and maintain its current growth.

  19. DNA Damage Response and Immune Defence: Links and Mechanisms

    Directory of Open Access Journals (Sweden)

    Björn Schumacher

    2016-08-01

    Full Text Available DNA damage plays a causal role in numerous human pathologies including cancer, premature aging and chronic inflammatory conditions. In response to genotoxic insults, the DNA damage response (DDR orchestrates DNA damage checkpoint activation and facilitates the removal of DNA lesions. The DDR can also arouse the immune system by for example inducing the expression of antimicrobial peptides as well as ligands for receptors found on immune cells. The activation of immune signalling is triggered by different components of the DDR including DNA damage sensors, transducer kinases, and effectors. In this review, we describe recent advances on the understanding of the role of DDR in activating immune signalling. We highlight evidence gained into (i which molecular and cellular pathways of DDR activate immune signalling, (ii how DNA damage drives chronic inflammation, and (iii how chronic inflammation causes DNA damage and pathology in humans.

  20. Beyond CTLA-4 and PD-1, the generation Z of negative checkpoint regulators.

    Directory of Open Access Journals (Sweden)

    Isabelle eLe Mercier

    2015-08-01

    Full Text Available In the last two years, clinical trials with blocking antibodies to the negative checkpoint regulators CTLA-4 and PD-1 have rekindled the hope for cancer immunotherapy. Multiple negative checkpoint regulators protect the host against autoimmune reactions but also restrict the ability of T cells to effectively attack tumors. Releasing these brakes has emerged as an exciting strategy for cancer treatment. Conversely, these pathways can be manipulated to achieve durable tolerance for treatment of autoimmune diseases and transplantation. In the future, treatment may involve combination therapy to target multiple cell types and stages of the adaptive immune responses. In this review, we describe the current knowledge on the recently discovered negative checkpoint regulators, future targets for immunotherapy.

  1. Beyond CTLA-4 and PD-1, the Generation Z of Negative Checkpoint Regulators.

    Science.gov (United States)

    Le Mercier, Isabelle; Lines, J Louise; Noelle, Randolph J

    2015-01-01

    In the last two years, clinical trials with blocking antibodies to the negative checkpoint regulators CTLA-4 and PD-1 have rekindled the hope for cancer immunotherapy. Multiple negative checkpoint regulators protect the host against autoimmune reactions but also restrict the ability of T cells to effectively attack tumors. Releasing these brakes has emerged as an exciting strategy for cancer treatment. Conversely, these pathways can be manipulated to achieve durable tolerance for treatment of autoimmune diseases and transplantation. In the future, treatment may involve combination therapy to target multiple cell types and stages of the adaptive immune responses. In this review, we describe the current knowledge on the recently discovered negative checkpoint regulators, future targets for immunotherapy.

  2. A tumor suppressor role of the Bub3 spindle checkpoint protein after apoptosis inhibition

    Science.gov (United States)

    Moutinho-Santos, Tatiana

    2013-01-01

    Most solid tumors contain aneuploid cells, indicating that the mitotic checkpoint is permissive to the proliferation of chromosomally aberrant cells. However, mutated or altered expression of mitotic checkpoint genes accounts for a minor proportion of human tumors. We describe a Drosophila melanogaster tumorigenesis model derived from knocking down spindle assembly checkpoint (SAC) genes and preventing apoptosis in wing imaginal discs. Bub3-deficient tumors that were also deficient in apoptosis displayed neoplastic growth, chromosomal aneuploidy, and high proliferative potential after transplantation into adult flies. Inducing aneuploidy by knocking down CENP-E and preventing apoptosis does not induce tumorigenesis, indicating that aneuploidy is not sufficient for hyperplasia. In this system, the aneuploidy caused by a deficient SAC is not driving tumorigenesis because preventing Bub3 from binding to the kinetochore does not cause hyperproliferation. Our data suggest that Bub3 has a nonkinetochore-dependent function that is consistent with its role as a tumor suppressor. PMID:23609535

  3. Use of common time base for checkpointing and rollback recovery in a distributed system

    Science.gov (United States)

    Ramanathan, Parameswaran; Shin, Kang G.

    1993-01-01

    An approach to checkpointing and rollback recovery in a distributed computing system using a common time base is proposed. A common time base is established in the system using a hardware clock synchronization algorithm. This common time base is coupled with the idea of pseudo-recovery points to develop a checkpointing algorithm that has the following advantages: reduced wait for commitment for establishing recovery lines, fewer messages to be exchanged, and less memory requirement. These advantages are assessed quantitatively by developing a probabilistic model.

  4. Cyclin F suppresses B-Myb activity to promote cell cycle checkpoint control

    DEFF Research Database (Denmark)

    Klein, Ditte Kjærsgaard; Hoffmann, Saskia; Ahlskog, Johanna K

    2015-01-01

    an important role in checkpoint control following ionizing radiation. Cyclin F-depleted cells initiate checkpoint signalling after ionizing radiation, but fail to maintain G2 phase arrest and progress into mitosis prematurely. Importantly, cyclin F suppresses the B-Myb-driven transcriptional programme...... that promotes accumulation of crucial mitosis-promoting proteins. Cyclin F interacts with B-Myb via the cyclin box domain. This interaction is important to suppress cyclin A-mediated phosphorylation of B-Myb, a key step in B-Myb activation. In summary, we uncover a regulatory mechanism linking the F-box protein...

  5. Interprofessional Collaboration with Immune Checkpoint Inhibitor Therapy: the Roles of Gastroenterology, Endocrinology and Neurology.

    Science.gov (United States)

    Seery, Virginia

    2017-11-01

    To discuss immune checkpoint inhibitor therapy and identify opportunities for interprofessional collaboration in the management of toxicities in the areas of gastroenterology, endocrinology, and neurology. Published research and education articles in oncology, nursing, and various specialties. The use of immune checkpoint inhibitors is expanding; timely management of toxicity is critical for positive patient outcomes. There are many opportunities for interprofessional collaboration in the diagnosis and treatment of immune-related adverse events. Nurses play key roles in recognizing immune-related adverse events, providing patient education, and helping to facilitate interprofessional collaboration. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Immune Checkpoint Function of CD85j in CD8 T Cell Differentiation and Aging

    Directory of Open Access Journals (Sweden)

    Claire E. Gustafson

    2017-06-01

    Full Text Available Aging is associated with an increased susceptibility to infection and a failure to control latent viruses thought to be driven, at least in part, by alterations in CD8 T cell function. The aging T cell repertoire is characterized by an accumulation of effector CD8 T cells, many of which express the negative regulatory receptor CD85j. To define the biological significance of CD85j expression on CD8 T cells and to address the question whether presence of CD85j in older individuals is beneficial or detrimental for immune function, we examined the specific attributes of CD8 T cells expressing CD85j as well as the functional role of CD85j in antigen-specific CD8 T cell responses during immune aging. Here, we show that CD85j is mainly expressed by terminally differentiated effector (TEMRAs CD8 T cells, which increase with age, in cytomegalovirus (CMV infection and in males. CD85j+ CMV-specific cells demonstrate clonal expansion. However, TCR diversity is similar between CD85j+ and CD85j− compartments, suggesting that CD85j does not directly impact the repertoire of antigen-specific cells. Further phenotypic and functional analyses revealed that CD85j identifies a specific subset of CMV-responsive CD8 T cells that coexpress a marker of senescence (CD57 but retain polyfunctional cytokine production and expression of cytotoxic mediators. Blocking CD85j binding enhanced proliferation of CMV-specific CD8 T cells upon antigen stimulation but did not alter polyfunctional cytokine production. Taken together, these data demonstrate that CD85j characterizes a population of “senescent,” but not exhausted antigen-specific effector CD8 T cells and indicates that CD85j is an important checkpoint regulator controlling expansion of virus-specific T cells during aging. Inhibition of CD85j activity may be a mechanism to promote stronger CD8 T cell effector responses during immune aging.

  7. Molecular Checkpoint Decisions Made by Subverted Vascular Niche Transform Indolent Tumor Cells into Chemoresistant Cancer Stem Cells.

    Science.gov (United States)

    Cao, Zhongwei; Scandura, Joseph M; Inghirami, Giorgio G; Shido, Koji; Ding, Bi-Sen; Rafii, Shahin

    2017-01-09

    Tumor-associated endothelial cells (TECs) regulate tumor cell aggressiveness. However, the core mechanism by which TECs confer stem cell-like activity to indolent tumors is unknown. Here, we used in vivo murine and human tumor models to identify the tumor-suppressive checkpoint role of TEC-expressed insulin growth factor (IGF) binding protein-7 (IGFBP7/angiomodulin). During tumorigenesis, IGFBP7 blocks IGF1 and inhibits expansion and aggresiveness of tumor stem-like cells (TSCs) expressing IGF1 receptor (IGF1R). However, chemotherapy triggers TECs to suppress IGFBP7, and this stimulates IGF1R + TSCs to express FGF4, inducing a feedforward FGFR1-ETS2 angiocrine cascade that obviates TEC IGFBP7. Thus, loss of IGFBP7 and upregulation of IGF1 activates the FGF4-FGFR1-ETS2 pathway in TECs and converts naive tumor cells to chemoresistant TSCs, thereby facilitating their invasiveness and progression. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Genetic variation in the major mitotic checkpoint genes associated with chromosomal aberrations in healthy humans

    Czech Academy of Sciences Publication Activity Database

    Försti, A.; Frank, Ch.; Smolková, B.; Kazimírová, A.; Barančoková, M.; Vymetálková, Veronika; Kroupa, M.; Naccarati, Alessio; Vodičková, Ludmila; Buchancová, J.; Dusinská, M.; Musak, L.; Vodička, Pavel; Hemminki, K.

    2016-01-01

    Roč. 380, č. 2 (2016), s. 442-446 ISSN 0304-3835 R&D Projects: GA ČR(CZ) GA15-14789S Institutional support: RVO:68378041 Keywords : chromosomal integrity * cytogenetics * spindle checkpoint Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.375, year: 2016

  9. Synthesis of Fault-Tolerant Embedded Systems with Checkpointing and Replication

    DEFF Research Database (Denmark)

    Izosimov, Viacheslav; Pop, Paul; Eles, Petru

    2006-01-01

    We present an approach to the synthesis of fault-tolerant hard real-time systems for safety-critical applications. We use checkpointing with rollback recovery and active replication for tolerating transient faults. Processes are statically scheduled and communications are performed using the time...

  10. Constitutive Cdk2 activity promotes aneuploidy while altering the spindle assembly and tetraploidy checkpoints

    DEFF Research Database (Denmark)

    Jahn, Stephan C; Corsino, Patrick E; Davis, Bradley J

    2013-01-01

    instability. Expression of these complexes in the MCF10A cell line leads to retinoblastoma protein (Rb) hyperphosphorylation, a subsequent increase in proliferation rate, and increased expression of the spindle assembly checkpoint protein Mad2. This results in a strengthening of the spindle assembly...

  11. Localization of spindle checkpoint proteins in cells undergoing mitosis with unreplicated genomes.

    Science.gov (United States)

    Johnson, Mary Kathrine; Cooksey, Amanda M; Wise, Dwayne A

    2008-11-01

    CHO cells can be arrested with hydoxyurea at the beginning of the DNA synthesis phase of the cell cycle. Subsequent treatment with the xanthine, caffeine, induces cells to bypass the S-phase checkpoint and enter unscheduled mitosis [Schlegel and Pardee,1986, Science 232:1264-1266]. These treated cells build a normal spindle and distribute kinetochores, unattached to chromosomes, to their daughter cells [Brinkley et al.,1988, Nature 336:251-254; Zinkowski et al.,1991, J Cell Biol 113:1091-1110; Wise and Brinkley,1997, Cell Motil Cytoskeleton 36:291-302; Balczon et al.,2003, Chromosoma 112:96-102]. To investigate how these cells distribute kinetochores to daughter cells, we analyzed the spindle checkpoint components, Mad2, CENP-E, and the 3F3 phosphoepitope, using immunofluorescence and digital microscopy. Even though the kinetochores were unpaired and DNA was fragmented, the tension, alignment, and motor components of the checkpoint were found to be present and localized as predicted in prometaphase and metaphase. This unusual mitosis proves that a cell can successfully localize checkpoint proteins and divide even when kinetochores are unpaired and fragmented. (c) 2008 Wiley-Liss, Inc.

  12. The point of no return: The poly(A)-associated elongation checkpoint.

    Science.gov (United States)

    Tellier, Michael; Ferrer-Vicens, Ivan; Murphy, Shona

    2016-01-01

    Cyclin-dependent kinases play critical roles in transcription by RNA polymerase II (pol II) and processing of the transcripts. For example, CDK9 regulates transcription of protein-coding genes, splicing, and 3' end formation of the transcripts. Accordingly, CDK9 inhibitors have a drastic effect on the production of mRNA in human cells. Recent analyses indicate that CDK9 regulates transcription at the early-elongation checkpoint of the vast majority of pol II-transcribed genes. Our recent discovery of an additional CDK9-regulated elongation checkpoint close to poly(A) sites adds a new layer to the control of transcription by this critical cellular kinase. This novel poly(A)-associated checkpoint has the potential to powerfully regulate gene expression just before a functional polyadenylated mRNA is produced: the point of no return. However, many questions remain to be answered before the role of this checkpoint becomes clear. Here we speculate on the possible biological significance of this novel mechanism of gene regulation and the players that may be involved.

  13. The point of no return: The poly(A)-associated elongation checkpoint

    Science.gov (United States)

    Tellier, Michael; Ferrer-Vicens, Ivan; Murphy, Shona

    2016-01-01

    abstract Cyclin-dependent kinases play critical roles in transcription by RNA polymerase II (pol II) and processing of the transcripts. For example, CDK9 regulates transcription of protein-coding genes, splicing, and 3′ end formation of the transcripts. Accordingly, CDK9 inhibitors have a drastic effect on the production of mRNA in human cells. Recent analyses indicate that CDK9 regulates transcription at the early-elongation checkpoint of the vast majority of pol II-transcribed genes. Our recent discovery of an additional CDK9-regulated elongation checkpoint close to poly(A) sites adds a new layer to the control of transcription by this critical cellular kinase. This novel poly(A)-associated checkpoint has the potential to powerfully regulate gene expression just before a functional polyadenylated mRNA is produced: the point of no return. However, many questions remain to be answered before the role of this checkpoint becomes clear. Here we speculate on the possible biological significance of this novel mechanism of gene regulation and the players that may be involved. PMID:26853452

  14. Immune Checkpoint Inhibitors in the Treatment of Patients with Neuroendocrine Neoplasia.

    Science.gov (United States)

    Weber, Matthias M; Fottner, Christian

    2018-01-01

    Well-differentiated neuroendocrine neoplasms (NENs) are usually controlled by antiproliferative, local ablative and/or radionuclide therapies, whereas poorly differentiated NENs generally require cytotoxic chemotherapy. However, treatment options for patients with advanced/metastatic high-grade NENs remain limited. Review of the literature and international congress abstracts on the efficacy and safety of immunotherapy by checkpoint inhibition in advanced/metastatic NENs. Evidence points to an important role of immune phenomena in the pathogenesis and treatment of neuroendocrine tumors (NETs). Programmed cell death 1 (PD-1) protein and its ligand are mainly expressed in poorly differentiated NENs. Microsatellite instability and high mutational load are more pronounced in high-grade NENs and may predict response to immunotherapy. Clinical experience of immune checkpoint blockade mainly exists for Merkel cell carcinoma, a high-grade cutaneous neuroendocrine carcinoma (NEC), which has led to approval of the anti-PD-1 antibody avelumab. In addition, there is anecdotal evidence for the efficacy of checkpoint inhibitors in large-cell lung NECs, ovarian NECs and others, including gastroenteropancreatic NENs. Currently, phase II studies investigate PDR001, pembrolizumab, combined durvalumab and tremelimumab, and avelumab treatment in patients with advanced/metastatic NENs. Immune checkpoint inhibitors are a promising therapeutic option, especially in progressive NECs or high-grade NETs with high tumor burden, microsatellite instability, and/or mutational load. © 2018 S. Karger GmbH, Freiburg.

  15. Myasthenia triggered by immune checkpoint inhibitors: New case and literature review.

    Science.gov (United States)

    Gonzalez, Natalia L; Puwanant, Araya; Lu, Angela; Marks, Stanley M; Živković, Saša A

    2017-03-01

    Immune checkpoint molecules are potent regulators of immunologic homeostasis that prevent the development of autoimmunity while maintaining self-tolerance. Inhibitors of immune checkpoint molecules are used as immunotherapy in the treatment of melanoma and different types of refractory cancer, and can trigger various autoimmune complications including myositis and myasthenia gravis. We describe a case of generalized myasthenia gravis induced by pembrolizumab and review 11 other cases. Five patients also had elevated serum CK levels ranging from 1200 to 8729 IU/L, and biopsy showed myositis in one. Severity was highly variable as symptoms normalized spontaneously in one patient, but three others developed myasthenic crisis (including two with fatal outcomes). Steroids have been recommended as a preferred treatment of autoimmune complications of immune-checkpoint inhibitors. Myasthenia gravis should be considered when weakness, diplopia or bulbar symptoms are seen after treatment with immune checkpoint inhibitors, and additional studies are needed to characterize association with hyperCKemia. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Immunogenic Chemotherapy Sensitizes Renal Cancer to Immune Checkpoint Blockade Therapy in Preclinical Models.

    Science.gov (United States)

    Cui, Shujin

    2017-07-11

    BACKGROUND Renal cell carcinoma (RCC) is among the most common malignant cancers of males worldwide. For advanced RCC patients, there still is no effective therapy. Immune checkpoint blockade therapies have shown benefits for many cancers, but previous clinical trials of immune checkpoint blockade therapies in RCC patients achieved only modest results. MATERIAL AND METHODS We explored the effects of combining chemotherapy with immune checkpoint blockade therapy in RCC xenograft mouse models. We also studied the potential mechanisms by which chemotherapy might enhance the efficacy of immune checkpoint blockade therapy, both in vitro and in vivo. RESULTS Our results showed that many commonly used chemotherapy agents can induce immunogenic marker release in RCC cell lines. Importantly, the RCC xenograft mouse model mice who received the combination treatment of 5-fluorouracil (5-FU) and anti-programmed cell death-ligand 1 (PD-L1) antibodies (Abs) had longer survival times compared to those who received 5-FU or anti-PD-L1 Abs alone. Also, increased key cytokines that promote tumor immunity, such as IL-2, IFN-γ, and TNF-α, as well as tumor-infiltrating cytotoxic T cells, were also increased after the combination treatment. CONCLUSIONS We conclude that 5-FU can sensitize RCC to anti-PD-L1 treatment by releasing the immune suppression in the tumor microenvironment.

  17. The SFP1 gene product of Saccharomyces cerevisiae regulates G2/M transitions during the mitotic cell cycle and DNA-damage response

    International Nuclear Information System (INIS)

    Xu, Z.; Norris, D.

    1998-01-01

    In eukaryotic cells, checkpoint pathways arrest cell-cycle progression if a particular event has failed to complete appropriately or if an important intracellular structure is defective or damaged. Saccharomyces cerevisiae strains that lack the SFP1 gene fail to arrest at the G2 DNA-damage checkpoint in response to genomic injury, but maintain their ability to arrest at the replication and spindle-assembly checkpoints. sfp1D mutants are characterized by a premature entrance into mitosis during a normal (undamaged) cell cycle, while strains that overexpress Sfp1p exhibit delays in G2. Sfp1p therefore acts as a repressor of the G2/M transition, both in the normal cell cycle and in the G2 checkpoint pathway. Sfp1 is a nuclear protein with two Cys2His2 zinc-finger domains commonly found in transcription factors. We propose that Sfp1p regulates the expression of gene products involved in the G2/M transition during the mitotic cell cycle and the DNA-damage response. In support of this model, overexpression of Sfp1p induces the expression of the PDS1 gene, which is known to encode a protein that regulates the G2 checkpoint. (author)

  18. Leptin Regulation of Gonadotrope Gonadotropin-Releasing Hormone Receptors As a Metabolic Checkpoint and Gateway to Reproductive Competence

    Directory of Open Access Journals (Sweden)

    Angela K. Odle

    2018-01-01

    Full Text Available The adipokine leptin signals the body’s nutritional status to the brain, and particularly, the hypothalamus. However, leptin receptors (LEPRs can be found all throughout the body and brain, including the pituitary. It is known that leptin is permissive for reproduction, and mice that cannot produce leptin (Lep/Lep are infertile. Many studies have pinpointed leptin’s regulation of reproduction to the hypothalamus. However, LEPRs exist at all levels of the hypothalamic–pituitary–gonadal axis. We have previously shown that deleting the signaling portion of the LEPR specifically in gonadotropes impairs fertility in female mice. Our recent studies have targeted this regulation to the control of gonadotropin releasing hormone receptor (GnRHR expression. The hypotheses presented here are twofold: (1 cyclic regulation of pituitary GnRHR levels sets up a target metabolic checkpoint for control of the reproductive axis and (2 multiple checkpoints are required for the metabolic signaling that regulates the reproductive axis. Here, we emphasize and explore the relationship between the hypothalamus and the pituitary with regard to the regulation of GnRHR. The original data we present strengthen these hypotheses and build on our previous studies. We show that we can cause infertility in 70% of female mice by deleting all isoforms of LEPR specifically in gonadotropes. Our findings implicate activin subunit (InhBa mRNA as a potential leptin target in gonadotropes. We further show gonadotrope-specific upregulation of GnRHR protein (but not mRNA levels following leptin stimulation. In order to try and understand this post-transcriptional regulation, we tested candidate miRNAs (identified with in silico analysis that may be binding the Gnrhr mRNA. We show significant upregulation of one of these miRNAs in our gonadotrope-Lepr-null females. The evidence provided here, combined with our previous work, lay the foundation for metabolically regulated post

  19. The emerging role of immune checkpoint based approaches in AML and MDS.

    Science.gov (United States)

    Boddu, Prajwal; Kantarjian, Hagop; Garcia-Manero, Guillermo; Allison, James; Sharma, Padmanee; Daver, Naval

    2018-04-01

    The development of immune checkpoint inhibitors represents a major breakthrough in the field of cancer therapeutics. Pursuant to their success in melanoma and numerous solid tumor malignancies, these agents are being investigated in hematological malignancies including acute myelogenous leukemia (AML) and myelodysplastic syndromes (MDS). Although AML/MDS have traditionally been considered to be less immunogenic than solid tumor malignancies, recent pre-clinical models suggest a therapeutic role for immune checkpoint inhibition in these diseases. CTLA-4 inhibition may be especially effective in treating late post-allogeneic stem cell transplant relapse of AML in patients with limited or no graft versus host disease. Immune checkpoint inhibition, specifically PD-1 inhibition, demonstrated limited single agent efficacy in patients with relapsed AML and with MDS post-hypomethylating therapy. Rationally designed combinations of PD-1 inhibitors with standard anti-leukemic therapy are needed. Hypomethylating agents such as azacitidine, up-regulate PD-1, PD-L1, and PD-L2 in patients with AML/MDS and up-regulation of these genes was associated with the emergence of resistance. The combination of azacitidine and PD-1/PD-L1 inhibition may be a potential mechanism to prevent or overcome resistance to 5-azacitidine. A number of such combinations are being evaluated in clinical trials with early encouraging results. Immune checkpoint inhibition is also an attractive option to improve relapse-free survival or eliminate minimal residual disease post induction and consolidation by enhancing T-cell surveillance in patients with high-risk AML. The ongoing clinical trials with checkpoint inhibitors in AML/MDS will improve our understanding of the immunobiology of these diseases and guide us to the most appropriate application of these agents in the therapy of AML/MDS.

  20. New elements of molecular orchestra at radiation-induced damaged genomic sites

    International Nuclear Information System (INIS)

    Wani, Altaf A.; Battu, Aruna; Ray, Alo

    2012-01-01

    DNA damage promptly activates cell cycle checkpoints enabling cells to repair their genome. ATR and ATM kinases are central to the checkpoint activation in response to DNA damage and replication stress. Activated ATR and ATM phosphorylate several downstream proteins involved in DNA repair and cell cycle arrest. However, the nature of the signal, which initially activates these kinases in response to UV damage and how they interact with nucleotide excision repair (NER) pathway, is unclear. Our research has shown that DDB2 and XPC, two early damage recognition factors, promoted ATR and ATM recruitment and phosphorylation. ATR and ATM localized to the damage site and physically interact with XPC. ATR and ATM recruitment and their phosphorylation is negatively affected in cells defective in DDB2 and XPC functions while conversely ATR- and ATM-deficiency fail to influence the damage recruitment of DDB2 and XPC proteins. Consequently, the phosphorylation of ATR and ATM substrates, Chk1, Chk2, H2AX, and BRCA1 is significantly reduced or abrogated in mutant cells, indicating that defective DDB2 and XPC function impaired the checkpoint signal transduction cascade in response to UV damage. DDB2 and XPC also regulated the BRCA1 and Rad51 recruitment to the damage site, implicating their role in homologous recombination-mediated DNA repair pathway. Supporting data reveal that the depletion of ATR and ATM influenced the NER efficiency. Moreover, upon completion of NER, ordered restoration of chromatin structure and key epigenetic marks are necessary for resumption of the cell's normal function. We have demonstrated such a restoration role of H3K56 acetylation (H3K56Ac) mark in response to UV irradiation, In human cells a fast initial deacetylation of H3K56 is followed by full renewal of an acetylated state at ∼ 24-48 hr post-irradiation. Histone chaperone, anti-silencing function-1A (ASF1A), is crucial for post-repair H3K56Ac restoration, which in turn, is needed for the

  1. Caenorhabditis elegans histone methyltransferase MET-2 shields the male X chromosome from checkpoint machinery and mediates meiotic sex chromosome inactivation.

    Directory of Open Access Journals (Sweden)

    Paula M Checchi

    2011-09-01

    Full Text Available Meiosis is a specialized form of cellular division that results in the precise halving of the genome to produce gametes for sexual reproduction. Checkpoints function during meiosis to detect errors and subsequently to activate a signaling cascade that prevents the formation of aneuploid gametes. Indeed, asynapsis of a homologous chromosome pair elicits a checkpoint response that can in turn trigger germline apoptosis. In a heterogametic germ line, however, sex chromosomes proceed through meiosis with unsynapsed regions and are not recognized by checkpoint machinery. We conducted a directed RNAi screen in Caenorhabditis elegans to identify regulatory factors that prevent recognition of heteromorphic sex chromosomes as unpaired and uncovered a role for the SET domain histone H3 lysine 9 histone methyltransferase (HMTase MET-2 and two additional HMTases in shielding the male X from checkpoint machinery. We found that MET-2 also mediates the transcriptional silencing program of meiotic sex chromosome inactivation (MSCI but not meiotic silencing of unsynapsed chromatin (MSUC, suggesting that these processes are distinct. Further, MSCI and checkpoint shielding can be uncoupled, as double-strand breaks targeted to an unpaired, transcriptionally silenced extra-chromosomal array induce checkpoint activation in germ lines depleted for met-2. In summary, our data uncover a mechanism by which repressive chromatin architecture enables checkpoint proteins to distinguish between the partnerless male X chromosome and asynapsed chromosomes thereby shielding the lone X from inappropriate activation of an apoptotic program.

  2. The Pch2 AAA+ ATPase promotes phosphorylation of the Hop1 meiotic checkpoint adaptor in response to synaptonemal complex defects.

    Science.gov (United States)

    Herruzo, Esther; Ontoso, David; González-Arranz, Sara; Cavero, Santiago; Lechuga, Ana; San-Segundo, Pedro A

    2016-09-19

    Meiotic cells possess surveillance mechanisms that monitor critical events such as recombination and chromosome synapsis. Meiotic defects resulting from the absence of the synaptonemal complex component Zip1 activate a meiosis-specific checkpoint network resulting in delayed or arrested meiotic progression. Pch2 is an evolutionarily conserved AAA+ ATPase required for the checkpoint-induced meiotic block in the zip1 mutant, where Pch2 is only detectable at the ribosomal DNA array (nucleolus). We describe here that high levels of the Hop1 protein, a checkpoint adaptor that localizes to chromosome axes, suppress the checkpoint defect of a zip1 pch2 mutant restoring Mek1 activity and meiotic cell cycle delay. We demonstrate that the critical role of Pch2 in this synapsis checkpoint is to sustain Mec1-dependent phosphorylation of Hop1 at threonine 318. We also show that the ATPase activity of Pch2 is essential for its checkpoint function and that ATP binding to Pch2 is required for its localization. Previous work has shown that Pch2 negatively regulates Hop1 chromosome abundance during unchallenged meiosis. Based on our results, we propose that, under checkpoint-inducing conditions, Pch2 also possesses a positive action on Hop1 promoting its phosphorylation and its proper distribution on unsynapsed chromosome axes. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  3. Improvement of Antitumor Therapies Based on Vaccines and Immune-Checkpoint Inhibitors by Counteracting Tumor-Immunostimulation

    Directory of Open Access Journals (Sweden)

    Paula Chiarella

    2018-01-01

    Full Text Available Immune-checkpoint inhibitors and antitumor vaccines may produce both tumor-inhibitory and tumor-stimulatory effects on growing tumors depending on the stage of tumor growth at which treatment is initiated. These paradoxical results are not necessarily incompatible with current tumor immunology but they might better be explained assuming the involvement of the phenomenon of tumor immunostimulation. This phenomenon was originally postulated on the basis that the immune response (IR evoked in Winn tests by strong chemical murine tumors was not linear but biphasic, with strong IR producing inhibition and weak IR inducing stimulation of tumor growth. Herein, we extended those former observations to weak spontaneous murine tumors growing in pre-immunized, immune-competent and immune-depressed mice. Furthermore, we demonstrated that the interaction of specifical T cells and target tumor cells at low stimulatory ratios enhanced the production of chemokines aimed to recruit macrophages at the tumor site, which, upon activation of toll-like receptor 4 and p38 signaling pathways, would recruit and activate more macrophages and other inflammatory cells which would produce growth-stimulating signals leading to an accelerated tumor growth. On this basis, the paradoxical effects achieved by immunological therapies on growing tumors could be explained depending upon where the therapy-induced IR stands on the biphasic IR curve at each stage of tumor growth. At stages where tumor growth was enhanced (medium and large-sized tumors, counteraction of the tumor-immunostimulatory effect with anti-inflammatory strategies or, more efficiently, with selective inhibitors of p38 signaling pathways enabled the otherwise tumor-promoting immunological strategies to produce significant inhibition of tumor growth.

  4. Lovastatin prevents cisplatin-induced activation of pro-apoptotic DNA damage response (DDR) of renal tubular epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Krüger, Katharina; Ziegler, Verena; Hartmann, Christina; Henninger, Christian [Institute of Toxicology, Medical Faculty, Heinrich Heine University Düsseldorf, 40225 Düsseldorf (Germany); Thomale, Jürgen [Institute of Cell Biology, University Duisburg-Essen, 45122 Essen (Germany); Schupp, Nicole [Institute of Toxicology, Medical Faculty, Heinrich Heine University Düsseldorf, 40225 Düsseldorf (Germany); Fritz, Gerhard, E-mail: fritz@uni-duesseldorf.de [Institute of Toxicology, Medical Faculty, Heinrich Heine University Düsseldorf, 40225 Düsseldorf (Germany)

    2016-02-01

    The platinating agent cisplatin (CisPt) is commonly used in the therapy of various types of solid tumors. The anticancer efficacy of CisPt largely depends on the formation of bivalent DNA intrastrand crosslinks, which stimulate mechanisms of the DNA damage response (DDR), thereby triggering checkpoint activation, gene expression and cell death. The clinically most relevant adverse effect associated with CisPt treatment is nephrotoxicity that results from damage to renal tubular epithelial cells. Here, we addressed the question whether the HMG-CoA-reductase inhibitor lovastatin affects the DDR of renal cells by employing rat renal proximal tubular epithelial (NRK-52E) cells as in vitro model. The data show that lovastatin has extensive inhibitory effects on CisPt-stimulated DDR of NRK-52E cells as reflected on the levels of phosphorylated ATM, Chk1, Chk2, p53 and Kap1. Mitigation of CisPt-induced DDR by lovastatin was independent of the formation of DNA damage as demonstrated by (i) the analysis of Pt-(GpG) intrastrand crosslink formation by Southwestern blot analyses and (ii) the generation of DNA strand breaks as analyzed on the level of nuclear γH2AX foci and employing the alkaline comet assay. Lovastatin protected NRK-52E cells from the cytotoxicity of high CisPt doses as shown by measuring cell viability, cellular impedance and flow cytometry-based analyses of cell death. Importantly, the statin also reduced the level of kidney DNA damage and apoptosis triggered by CisPt treatment of mice. The data show that the lipid-lowering drug lovastatin extensively counteracts pro-apoptotic signal mechanisms of the DDR of tubular epithelial cells following CisPt injury. - Highlights: • Lovastatin blocks ATM/ATR-regulated DDR of tubular cells following CisPt treatment. • Lovastatin attenuates CisPt-induced activation of protein kinase ATM in vitro. • Statin-mediated DDR inhibition is independent of initial DNA damage formation. • Statin-mediated blockage of Cis

  5. Lovastatin prevents cisplatin-induced activation of pro-apoptotic DNA damage response (DDR) of renal tubular epithelial cells

    International Nuclear Information System (INIS)

    Krüger, Katharina; Ziegler, Verena; Hartmann, Christina; Henninger, Christian; Thomale, Jürgen; Schupp, Nicole; Fritz, Gerhard

    2016-01-01

    The platinating agent cisplatin (CisPt) is commonly used in the therapy of various types of solid tumors. The anticancer efficacy of CisPt largely depends on the formation of bivalent DNA intrastrand crosslinks, which stimulate mechanisms of the DNA damage response (DDR), thereby triggering checkpoint activation, gene expression and cell death. The clinically most relevant adverse effect associated with CisPt treatment is nephrotoxicity that results from damage to renal tubular epithelial cells. Here, we addressed the question whether the HMG-CoA-reductase inhibitor lovastatin affects the DDR of renal cells by employing rat renal proximal tubular epithelial (NRK-52E) cells as in vitro model. The data show that lovastatin has extensive inhibitory effects on CisPt-stimulated DDR of NRK-52E cells as reflected on the levels of phosphorylated ATM, Chk1, Chk2, p53 and Kap1. Mitigation of CisPt-induced DDR by lovastatin was independent of the formation of DNA damage as demonstrated by (i) the analysis of Pt-(GpG) intrastrand crosslink formation by Southwestern blot analyses and (ii) the generation of DNA strand breaks as analyzed on the level of nuclear γH2AX foci and employing the alkaline comet assay. Lovastatin protected NRK-52E cells from the cytotoxicity of high CisPt doses as shown by measuring cell viability, cellular impedance and flow cytometry-based analyses of cell death. Importantly, the statin also reduced the level of kidney DNA damage and apoptosis triggered by CisPt treatment of mice. The data show that the lipid-lowering drug lovastatin extensively counteracts pro-apoptotic signal mechanisms of the DDR of tubular epithelial cells following CisPt injury. - Highlights: • Lovastatin blocks ATM/ATR-regulated DDR of tubular cells following CisPt treatment. • Lovastatin attenuates CisPt-induced activation of protein kinase ATM in vitro. • Statin-mediated DDR inhibition is independent of initial DNA damage formation. • Statin-mediated blockage of Cis

  6. Piperine causes G1 phase cell cycle arrest and apoptosis in melanoma cells through checkpoint kinase-1 activation.

    Directory of Open Access Journals (Sweden)

    Neel M Fofaria

    Full Text Available In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR and checkpoint kinase 1 (Chk1. Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb. Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis.

  7. growth stimulant

    African Journals Online (AJOL)

    Effects of timing and duration of supplementation of LIVFIT VET ® (growth stimulant) as substitute for fish meal on the growth performance, haematology and clinical enzymes concentration of growing pigs.

  8. Casein kinase II is required for the spindle assembly checkpoint by regulating Mad2p in fission yeast

    Energy Technology Data Exchange (ETDEWEB)

    Shimada, Midori [Department of Biochemistry and Cell Biology, Graduate School of Medicine, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Yamamoto, Ayumu [Department of Chemistry, Shizuoka University, 836 Ohya, Suruga-ku, Sizuoka 422-8529 (Japan); Murakami-Tonami, Yuko; Nakanishi, Makoto; Yoshida, Takashi [Department of Biochemistry and Cell Biology, Graduate School of Medicine, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Aiba, Hirofumi [Laboratory of Molecular Microbiology, School of Agriculture, Nagoya University, Chikusa-ku, Nagoya 464-8601 (Japan); Murakami, Hiroshi, E-mail: hmura@med.nagoya-cu.ac.jp [Department of Biochemistry and Cell Biology, Graduate School of Medicine, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan)

    2009-10-23

    The spindle checkpoint is a surveillance mechanism that ensures the fidelity of chromosome segregation in mitosis. Here we show that fission yeast casein kinase II (CK2) is required for this checkpoint function. In the CK2 mutants mitosis occurs in the presence of a spindle defect, and the spindle checkpoint protein Mad2p fails to localize to unattached kinetochores. The CK2 mutants are sensitive to the microtubule depolymerising drug thiabendazole, which is counteracted by ectopic expression of mad2{sup +}. The level of Mad2p is low in the CK2 mutants. These results suggest that CK2 has a role in the spindle checkpoint by regulating Mad2p.

  9. MicroRNAs, the DNA damage response and cancer

    International Nuclear Information System (INIS)

    Wouters, Maikel D.; Gent, Dik C. van; Hoeijmakers, Jan H.J.; Pothof, Joris

    2011-01-01

    Many carcinogenic agents such as ultra-violet light from the sun and various natural and man-made chemicals act by damaging the DNA. To deal with these potentially detrimental effects of DNA damage, cells induce a complex DNA damage response (DDR) that includes DNA repair, cell cycle checkpoints, damage tolerance systems and apoptosis. This DDR is a potent barrier against carcinogenesis and defects within this response are observed in many, if not all, human tumors. DDR defects fuel the evolution of precancerous cells to malignant tumors, but can also induce sensitivity to DNA damaging agents in cancer cells, which can be therapeutically exploited by the use of DNA damaging treatment modalities. Regulation of and coordination between sub-pathways within the DDR is important for maintaining genome stability. Although regulation of the DDR has been extensively studied at the transcriptional and post-translational level, less is known about post-transcriptional gene regulation by microRNAs, the topic of this review. More specifically, we highlight current knowledge about DNA damage responsive microRNAs and microRNAs that regulate DNA damage response genes. We end by discussing the role of DNA damage response microRNAs in cancer etiology and sensitivity to ionizing radiation and other DNA damaging therapeutic agents.

  10. RNA interference regulates the cell cycle checkpoint through the RNA export factor, Ptr1, in fission yeast

    Energy Technology Data Exchange (ETDEWEB)

    Iida, Tetsushi, E-mail: tiida@nig.ac.jp [Division of Cytogenetics, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); The Graduate University for Advanced Studies, Sokendai, Mishima, 1111 Yata, Mishima 411-8540 (Japan); Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (JST), 4-1-8, Honcho, Kawaguchi-shi, Saitama 332-0012 (Japan); Iida, Naoko [Division of Mutagenesis, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); Tsutsui, Yasuhiro [Department of Life Science, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Nagatsuda-cho, Midori-ku, Yokohama 226-8501 (Japan); Yamao, Fumiaki [Division of Mutagenesis, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); The Graduate University for Advanced Studies, Sokendai, Mishima, 1111 Yata, Mishima 411-8540 (Japan); Kobayashi, Takehiko [Division of Cytogenetics, National Institute of Genetics, Mishima, 1111 Yata, Mishima 411-8540 (Japan); The Graduate University for Advanced Studies, Sokendai, Mishima, 1111 Yata, Mishima 411-8540 (Japan)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer RNAi is linked to the cell cycle checkpoint in fission yeast. Black-Right-Pointing-Pointer Ptr1 co-purifies with Ago1. Black-Right-Pointing-Pointer The ptr1-1 mutation impairs the checkpoint but does not affect gene silencing. Black-Right-Pointing-Pointer ago1{sup +} and ptr1{sup +} regulate the cell cycle checkpoint via the same pathway. Black-Right-Pointing-Pointer Mutations in ago1{sup +} and ptr1{sup +} lead to the nuclear accumulation of poly(A){sup +} RNAs. -- Abstract: Ago1, an effector protein of RNA interference (RNAi), regulates heterochromatin silencing and cell cycle arrest in fission yeast. However, the mechanism by which Ago1 controls cell cycle checkpoint following hydroxyurea (HU) treatment has not been elucidated. In this study, we show that Ago1 and other RNAi factors control cell cycle checkpoint following HU treatment via a mechanism independent of silencing. While silencing requires dcr1{sup +}, the overexpression of ago1{sup +} alleviated the cell cycle defect in dcr1{Delta}. Ago1 interacted with the mRNA export factor, Ptr1. The ptr1-1 mutation impaired cell cycle checkpoint but gene silencing was unaffected. Genetic analysis revealed that the regulation of cell cycle checkpoint by ago1{sup +} is dependent on ptr1{sup +}. Nuclear accumulation of poly(A){sup +} RNAs was detected in mutants of ago1{sup +} and ptr1{sup +}, suggesting there is a functional link between the cell cycle checkpoint and RNAi-mediated RNA quality control.

  11. Immune Checkpoint Inhibitors: A New Opportunity in the Treatment of Ovarian Cancer?

    Directory of Open Access Journals (Sweden)

    Gloria Mittica

    2016-07-01

    Full Text Available Epithelial ovarian cancer (EOC is the leading cause of death for gynecological cancer. The standard treatment for advanced stage is the combination of optimal debulking surgery and platinum-based chemotherapy. Nevertheless, recurrence is frequent (around 70% and prognosis is globally poor. New therapeutic agents are needed to improve survival. Since EOC is strongly immunogenic, immune checkpoint inhibitors are under evaluation for their capacity to contrast the “turn off” signals expressed by the tumor to escape the immune system and usually responsible for self-tolerance maintenance. This article reviews the literature on anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA-4, anti-PD-1, anti-PD-L1, and anti-PD-L2 antibodies in EOC and highlights their possible lines of development. Further studies are needed to better define the prognostic role of the immune checkpoint inhibitors, to identify predictors of response and the optimal clinical setting in EOC.

  12. Immune Checkpoint Inhibitors: An Innovation in Immunotherapy for the Treatment and Management of Patients with Cancer.

    Science.gov (United States)

    Dine, Jennifer; Gordon, RuthAnn; Shames, Yelena; Kasler, Mary Kate; Barton-Burke, Margaret

    2017-01-01

    Cancer survival rates are generally increasing in the United States. These trends have been partially attributed to improvement in therapeutic strategies. Cancer immunotherapy is an example of one of the newer strategies used to fight cancer, which primes or activates the immune system to produce antitumor effects. The first half of this review paper concisely describes the cell mechanisms that control antitumor immunity and the major immunotherapeutic strategies developed to target these mechanisms. The second half of the review discusses in greater depth immune checkpoint inhibitors that have recently demonstrated tremendous promise for the treatment of diverse solid tumor types, including melanoma, non-small cell lung cancer, and others. More specifically, the mechanisms of action, side effects, and patient and family management and education concerns are discussed to provide oncology nurses up-to-date information relevant to caring for cancer-affected patients treated with immune checkpoint inhibitors. Future directions for cancer immunotherapy are considered.

  13. The fission yeast spindle orientation checkpoint: a model that generates tension?

    Science.gov (United States)

    Gachet, Yannick; Reyes, Céline; Goldstone, Sherilyn; Tournier, Sylvie

    2006-10-15

    In all eukaryotes, the alignment of the mitotic spindle with the axis of cell polarity is essential for accurate chromosome segregation as well as for the establishment of cell fate, and thus morphogenesis, during development. Studies in invertebrates, higher eukaryotes and yeast suggest that astral microtubules interact with the cell cortex to position the spindle. These microtubules are thought to impose pushing or pulling forces on the spindle poles to affect the rotation or movement of the spindle. In the fission yeast model, where cell division is symmetrical, spindle rotation is dependent on the interaction of astral microtubules with the cortical actin cytoskeleton. In these cells, a bub1-dependent mitotic checkpoint, the spindle orientation checkpoint (SOC), is activated when the spindles fail to align with the cell polarity axis. In this paper we review the mechanism that orientates the spindle during mitosis in fission yeast, and discuss the consequences of misorientation on metaphase progression. Copyright 2006 John Wiley & Sons, Ltd.

  14. Cloud object store for checkpoints of high performance computing applications using decoupling middleware

    Science.gov (United States)

    Bent, John M.; Faibish, Sorin; Grider, Gary

    2016-04-19

    Cloud object storage is enabled for checkpoints of high performance computing applications using a middleware process. A plurality of files, such as checkpoint files, generated by a plurality of processes in a parallel computing system are stored by obtaining said plurality of files from said parallel computing system; converting said plurality of files to objects using a log structured file system middleware process; and providing said objects for storage in a cloud object storage system. The plurality of processes may run, for example, on a plurality of compute nodes. The log structured file system middleware process may be embodied, for example, as a Parallel Log-Structured File System (PLFS). The log structured file system middleware process optionally executes on a burst buffer node.

  15. A Comprehensive Review of US FDA-Approved Immune Checkpoint Inhibitors in Urothelial Carcinoma

    Directory of Open Access Journals (Sweden)

    Fu-Shun Hsu

    2017-01-01

    Full Text Available Few effective treatment options are available for patients with advanced or metastatic urothelial carcinoma (UC after unsuccessful first-line platinum-based chemotherapy. To date, immune checkpoint inhibitors are novel therapeutic agents for UC treatment. From May 2016 to May 2017, five anti-PD-1/PD-L1 monoclonal antibodies received accelerated or regular approval from the US Food and Drug Administration (FDA for the treatment of patients with locally advanced or metastatic UC. The present comprehensive review presents the background information of these five US FDA-approved anticancer agents to provide a basic but concise understanding of these agents for advanced studies. We summarize their immune checkpoint mechanisms, clinical efficacy, recommended usage protocols, adverse events, and the limitations of the PD-L1 biomarker assays.

  16. SUV2, which encodes an ATR-related cell cycle checkpoint and putative plant ATRIP, is required for aluminium-dependent root growth inhibition in Arabidopsis.

    Science.gov (United States)

    Sjogren, Caroline A; Larsen, Paul B

    2017-09-01

    A suppressor mutagenesis screen was conducted in order to identify second site mutations that could reverse the extreme hypersensitivity to aluminium (Al) seen for the Arabidopsis mutant, als3-1. From this screen, it was found that a loss-of-function mutation in the previously described SUV2 (SENSITIVE TO UV 2), which encodes a putative plant ATRIP homologue that is a component of the ATR-dependent cell checkpoint response, reversed the als3-1 phenotype. This included prevention of hallmarks associated with als3-1 including Al-dependent terminal differentiation of the root tip and transition to endoreduplication. From this analysis, SUV2 was determined to be required for halting cell cycle progression and triggering loss of the quiescent centre (QC) following exposure to Al. In conjunction with this, SUV2 was found to have a similar role as ATR, ALT2 and SOG1 in Al-dependent stoppage of root growth, all of which are required for promotion of expression of a suite of genes that likely are part of an Al-dependent DNA damage transcriptional response. This work argues that these Al response factors work together to detect Al-dependent damage and subsequently activate a DNA damage response pathway that halts the cell cycle and subsequently promotes QC differentiation and entrance into endocycling. © 2017 John Wiley & Sons Ltd.

  17. Dynein Light Intermediate Chain 2 Facilitates the Metaphase to Anaphase Transition by Inactivating the Spindle Assembly Checkpoint.

    Directory of Open Access Journals (Sweden)

    Sagar P Mahale

    Full Text Available The multi-functional molecular motor cytoplasmic dynein performs diverse essential roles during mitosis. The mechanistic importance of the dynein Light Intermediate Chain homologs, LIC1 and LIC2 is unappreciated, especially in the context of mitosis. LIC1 and LIC2 are believed to exist in distinct cytoplasmic dynein complexes as obligate subunits. LIC1 had earlier been reported to be required for metaphase to anaphase progression by inactivating the kinetochore-microtubule attachment-sensing arm of the spindle assembly checkpoint (SAC. However, the functional importance of LIC2 during mitosis remains elusive. Here we report prominent novel roles for the LIC2 subunit of cytoplasmic dynein in regulating the spindle assembly checkpoint. LIC2 depletion in mammalian cells led to prolonged metaphase arrest in the presence of an active SAC and also to stretched kinetochores, thus implicating it in SAC inactivation. Quantitative fluorescence microscopy of SAC components revealed accumulation of both attachment- and tension-sensing checkpoint proteins at metaphase kinetochores upon LIC2 depletion. These observations support a stronger and more diverse role in checkpoint inactivation for LIC2 in comparison to its close homolog LIC1. Our study uncovers a novel functional hierarchy during mitotic checkpoint inactivation between the closely related but homologous LIC subunits of cytoplasmic dynein. These subtle functional distinctions between dynein subpopulations could be exploited to study specific aspects of the spindle assembly checkpoint, which is a key mediator of fidelity in eukaryotic cell division.

  18. Disruption of spindle checkpoint function in rats following 28 days of repeated administration of renal carcinogens.

    Science.gov (United States)

    Kimura, Masayuki; Mizukami, Sayaka; Watanabe, Yousuke; Hasegawa-Baba, Yasuko; Onda, Nobuhiko; Yoshida, Toshinori; Shibutani, Makoto

    2016-02-01

    We previously reported that 28-day exposure to hepatocarcinogens that facilitate cell proliferation specifically alters the expression of G1/S checkpoint-related genes and proteins, induces aberrant early expression of ubiquitin D (UBD) at the G2 phase, and increases apoptosis in the rat liver, indicating G1/S and spindle checkpoint dysfunction. The present study aimed to determine the time of onset of carcinogen-specific cell-cycle disruption after repeated administration of renal carcinogens for up to 28 days. Rats were orally administered the renal carcinogens nitrofurantoin (NFT), 1-amino-2,4-dibromoantraquinone (ADAQ), and 1,2,3-trichloropropane (TCP) or the non-carcinogenic renal toxicants 1-chloro-2-propanol, triamterene, and carboxin for 3, 7 or 28 days. Both immunohistochemical single-molecule analysis and real-time RT-PCR analysis revealed that carcinogen-specific expression changes were not observed after 28 days of administration. However, the renal carcinogens ADAQ and TCP specifically reduced the number of cells expressing phosphorylated-histone H3 at Ser10 in both UBD(+) cells and proliferating cells, suggestive of insufficient UBD expression at the M phase and early transition of proliferating cells from the M phase, without increasing apoptosis, after 28 days of administration. In contrast, NFT, which has marginal carcinogenic potential, did not induce such cellular responses. These results suggest that it may take 28 days to induce spindle checkpoint dysfunction by renal carcinogens; however, induction of apoptosis may not be essential. Thus, induction of spindle checkpoint dysfunction may be dependent on carcinogenic potential of carcinogen examined, and marginal carcinogens may not exert sufficient responses even after 28 days of administration.

  19. SPARC: Demonstrate burst-buffer-based checkpoint/restart on ATS-1.

    Energy Technology Data Exchange (ETDEWEB)

    Oldfield, Ron A. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Ulmer, Craig D. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Widener, Patrick [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Ward, H. Lee [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2018-01-01

    Recent high-performance computing (HPC) platforms such as the Trinity Advanced Technology System (ATS-1) feature burst buffer resources that can have a dramatic impact on an application’s I/O performance. While these non-volatile memory (NVM) resources provide a new tier in the storage hierarchy, developers must find the right way to incorporate the technology into their applications in order to reap the benefits. Similar to other laboratories, Sandia is actively investigating ways in which these resources can be incorporated into our existing libraries and workflows without burdening our application developers with excessive, platform-specific details. This FY18Q1 milestone summaries our progress in adapting the Sandia Parallel Aerodynamics and Reentry Code (SPARC) in Sandia’s ATDM program to leverage Trinity’s burst buffers for checkpoint/restart operations. We investigated four different approaches with varying tradeoffs in this work: (1) simply updating job script to use stage-in/stage out burst buffer directives, (2) modifying SPARC to use LANL’s hierarchical I/O (HIO) library to store/retrieve checkpoints, (3) updating Sandia’s IOSS library to incorporate the burst buffer in all meshing I/O operations, and (4) modifying SPARC to use our Kelpie distributed memory library to store/retrieve checkpoints. Team members were successful in generating initial implementation for all four approaches, but were unable to obtain performance numbers in time for this report (reasons: initial problem sizes were not large enough to stress I/O, and SPARC refactor will require changes to our code). When we presented our work to the SPARC team, they expressed the most interest in the second and third approaches. The HIO work was favored because it is lightweight, unobtrusive, and should be portable to ATS-2. The IOSS work is seen as a long-term solution, and is favored because all I/O work (including checkpoints) can be deferred to a single library.

  20. A Review of Immune Checkpoint Inhibitors for the Management of Locally Advanced or Metastatic Urothelial Carcinoma.

    Science.gov (United States)

    Hanna, Kirollos S

    2017-11-01

    Urothelial carcinoma (UC) is the second most common malignancy of the genitourinary system and the sixth most common cancer in the United States. The overall incidence of UC appears to be on the decline, but death rates have remained stable. Stage IV metastatic disease is associated with only a 5% survival rate at 5 years. Gemcitabine and cisplatin combinations or dose-dense methotrexate, vinblastine, doxorubicin, and cisplatin are the preferred regimens for individuals with advance, metastatic disease and a good performance status and organ function. Second-line therapies in this setting are limited. During the course of 1 year, five immune checkpoint inhibitors were approved for treatment of cancers in the locally advanced or metastatic setting: atezolizumab, nivolumab, durvalumab, avelumab, and pembrolizumab. Immunotherapies have played a significant role in the treatment of various cancers and have continued to expand. It is of utmost importance that practitioners include checkpoint inhibitors as treatment options for UC. Based on the limited data, pembrolizumab and atezolizumab may be the drugs of choice, as they are supported by the most influential data to date; however, further research is warranted. Ongoing clinical trials will further assess the benefits of inducing cellular immunity in the treatment of UC. These therapies mark a new landscape in the treatment of UC. In this article, the available data on immune checkpoint inhibitors for the treatment of locally advanced or metastatic UC and their place in therapy are reviewed. © 2017 Pharmacotherapy Publications, Inc.

  1. Measures to detect and control radioactive contaminated metallurgical scrap at border checkpoints in Poland

    International Nuclear Information System (INIS)

    Smagala, G.

    1999-01-01

    The issue of radioactive contaminated metallurgical scrap has never received a high priority in Poland and in the international community. Since the dissolution of the former Soviet Union a higher attention has been given to the problem. Poland which is located between the West and East Europe has the obligation to develop and implement an effective prevention and detection system. The reasons to increase national control and detection system at the border checkpoints in Poland are to avoid the following risks: post Chernobyl contamination transports of commodities; transport of contaminated metal scrap; transfer of radioactive waste for their disposal or utilization; high risk of becoming a transit country of illicit trafficking of nuclear materials and radioactive sources. In order to avoid the above-mentioned risks, Poland initiated in 1990, a deployment of the portable radiation devices at the border checkpoints and, as of 1998, the number of installed instruments exceeded a hundred. This paper presents Poland's activities to detect contaminated scrap at its border checkpoints. (author)

  2. TAM receptor tyrosine kinases as emerging targets of innate immune checkpoint blockade for cancer therapy.

    Science.gov (United States)

    Akalu, Yemsratch T; Rothlin, Carla V; Ghosh, Sourav

    2017-03-01

    Cancer immunotherapy utilizing T-cell checkpoint inhibitors has shown tremendous clinical success. Yet, this mode of treatment is effective in only a subset of patients. Unresponsive patients tend to have non-T-cell-inflamed tumors that lack markers associated with the activation of adaptive anti-tumor immune responses. Notably, elimination of cancer cells by T cells is critically dependent on the optimal activity of innate immune cells. Therefore, identifying new targets that regulate innate immune cell function and promote the engagement of adaptive tumoricidal responses is likely to lead to the development of improved therapies against cancer. Here, we review the TAM receptor tyrosine kinases-TYRO3, AXL, and MERTK-as an emerging class of innate immune checkpoints that participate in key steps of anti-tumoral immunity. Namely, TAM-mediated efferocytosis, negative regulation of dendritic cell activity, and dysregulated production of chemokines collectively favor the escape of malignant cells. Hence, disabling TAM signaling may promote engagement of adaptive immunity and complement T-cell checkpoint blockade. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Unique cytologic features of thyroiditis caused by immune checkpoint inhibitor therapy for malignant melanoma

    Directory of Open Access Journals (Sweden)

    Trevor E. Angell

    2018-03-01

    Full Text Available Blockade of immune checkpoint molecules to reverse cancer-induced immune suppression can improve anti-tumor immune responses in cancer patients. Monoclonal antibodies targeting two such molecules, Programmed cell death protein 1 (PD-1 and cytotoxic T-lymphocyte associated protein 4 (CTLA-4 have shown clinical benefit in the treatment of advanced malignancies, including metastatic melanoma. Adverse effects of these immune checkpoint inhibitors include immune-related adverse events (irAE, of which one of the most common is autoimmune thyroiditis. Though thyroiditis is increasingly recognized, there are no reports of the pathological findings that occur in immunotherapy-induced thyroiditis. We present a case of immunotherapy-induced thyroiditis demonstrating its unique cytopathologic features. A 51-year-old woman with metastatic melanoma was found to have a suppressed TSH and elevated free thyroxine concentration 14 days after starting treatment with nivolumab (PD-1 antagonist plus ipilimumab (CTLA-4 antagonist therapy. A thyroid biopsy was performed based on ultrasound findings and cytopathology revealed unique features including abundant clusters of necrotic cells, lymphocytes and CD163-positive histiocytes. This case reports cytopathologic features found in immune checkpoint inhibitor related thyroiditis. These appear to be unique findings and may help inform future research regarding the pathophysiology and mechanisms of this condition.

  4. Immune checkpoint inhibitors: the new frontier in non–small cell lung cancer treatment

    Directory of Open Access Journals (Sweden)

    El-Osta HE

    2016-08-01

    Full Text Available Hazem El-Osta, Kamran Shahid, Glenn M Mills, Prakash Peddi Department of Medicine, Division of Hematology-Oncology, Louisiana State University Health Sciences Center, Shreveport, LA, USA Abstract: Lung cancer is the major cause for cancer-related death in the US. Although advances in chemotherapy and targeted therapy have improved the outcome of metastatic non-small-cell lung cancer, its prognosis remains dismal. A deeper understanding of the complex interaction between the immune system and tumor microenvironment has identified immune checkpoint inhibitors as new avenue of immunotherapy. Rather than acting directly on the tumor, these therapies work by removing the inhibition exerted by tumor cell or other immune cells on the immune system, promoting antitumoral immune response. To date, two programmed death-1 inhibitors, namely nivolumab and pembrolizumab, have received the US Food and Drug Administration approval for the treatment of advanced non-small-cell lung cancer that failed platinum-based chemotherapy. This manuscript provides a brief overview of the pathophysiology of cancer immune evasion, summarizes pertinent data on completed and ongoing clinical trials involving checkpoint inhibitors, discusses the different strategies to optimize their function, and outlines various challenges that are faced in this promising yet evolving field. Keywords: checkpoint inhibitors, immunotherapy, nivolumab, non-small-cell lung cancer, pembrolizumab, programmed death-1, programmed death ligand-1

  5. Viral oncogene-induced DNA damage response is activated in Kaposi sarcoma tumorigenesis.

    Directory of Open Access Journals (Sweden)

    Sonja Koopal

    2007-09-01

    Full Text Available Kaposi sarcoma is a tumor consisting of Kaposi sarcoma herpesvirus (KSHV-infected tumor cells that express endothelial cell (EC markers and viral genes like v-cyclin, vFLIP, and LANA. Despite a strong link between KSHV infection and certain neoplasms, de novo virus infection of human primary cells does not readily lead to cellular transformation. We have studied the consequences of expression of v-cyclin in primary and immortalized human dermal microvascular ECs. We show that v-cyclin, which is a homolog of cellular D-type cyclins, induces replicative stress in ECs, which leads to senescence and activation of the DNA damage response. We find that antiproliferative checkpoints are activated upon KSHV infection of ECs, and in early-stage but not late-stage lesions of clinical Kaposi sarcoma specimens. These are some of the first results suggesting that DNA damage checkpoint response also functions as an anticancer barrier in virally induced cancers.

  6. The Chromatin Scaffold Protein SAFB1 Renders Chromatin Permissive for DNA Damage Signaling

    DEFF Research Database (Denmark)

    Altmeyer, Matthias; Toledo Lazaro, Luis Ignacio; Gudjonsson, Thorkell

    2013-01-01

    Although the general relevance of chromatin modifications for genotoxic stress signaling, cell-cycle checkpoint activation, and DNA repair is well established, how these modifications reach initial thresholds in order to trigger robust responses remains largely unexplored. Here, we identify...... the chromatin-associated scaffold attachment factor SAFB1 as a component of the DNA damage response and show that SAFB1 cooperates with histone acetylation to allow for efficient γH2AX spreading and genotoxic stress signaling. SAFB1 undergoes a highly dynamic exchange at damaged chromatin in a poly......(ADP-ribose)-polymerase 1- and poly(ADP-ribose)-dependent manner and is required for unperturbed cell-cycle checkpoint activation and guarding cells against replicative stress. Altogether, our data reveal that transient recruitment of an architectural chromatin component is required in order to overcome physiological...

  7. ZTF-8 interacts with the 9-1-1 complex and is required for DNA damage response and double-strand break repair in the C. elegans germline.

    Directory of Open Access Journals (Sweden)

    Hyun-Min Kim

    2014-10-01

    Full Text Available Germline mutations in DNA repair genes are linked to tumor progression. Furthermore, failure in either activating a DNA damage checkpoint or repairing programmed meiotic double-strand breaks (DSBs can impair chromosome segregation. Therefore, understanding the molecular basis for DNA damage response (DDR and DSB repair (DSBR within the germline is highly important. Here we define ZTF-8, a previously uncharacterized protein conserved from worms to humans, as a novel factor involved in the repair of both mitotic and meiotic DSBs as well as in meiotic DNA damage checkpoint activation in the C. elegans germline. ztf-8 mutants exhibit specific sensitivity to γ-irradiation and hydroxyurea, mitotic nuclear arrest at S-phase accompanied by activation of the ATL-1 and CHK-1 DNA damage checkpoint kinases, as well as accumulation of both mitotic and meiotic recombination intermediates, indicating that ZTF-8 functions in DSBR. However, impaired meiotic DSBR progression partially fails to trigger the CEP-1/p53-dependent DNA damage checkpoint in late pachytene, also supporting a role for ZTF-8 in meiotic DDR. ZTF-8 partially co-localizes with the 9-1-1 DDR complex and interacts with MRT-2/Rad1, a component of this complex. The human RHINO protein rescues the phenotypes observed in ztf-8 mutants, suggesting functional conservation across species. We propose that ZTF-8 is involved in promoting repair at stalled replication forks and meiotic DSBs by transducing DNA damage checkpoint signaling via the 9-1-1 pathway. Our findings define a conserved function for ZTF-8/RHINO in promoting genomic stability in the germline.

  8. Phosphorylation of the Budding Yeast 9-1-1 Complex Is Required for Dpb11 Function in the Full Activation of the UV-Induced DNA Damage Checkpoint▿ †

    Science.gov (United States)

    Puddu, Fabio; Granata, Magda; Di Nola, Lisa; Balestrini, Alessia; Piergiovanni, Gabriele; Lazzaro, Federico; Giannattasio, Michele; Plevani, Paolo; Muzi-Falconi, Marco

    2008-01-01

    Following genotoxic insults, eukaryotic cells trigger a signal transduction cascade known as the DNA damage checkpoint response, which involves the loading onto DNA of an apical kinase and several downstream factors. Chromatin modifications play an important role in recruiting checkpoint proteins. In budding yeast, methylated H3-K79 is bound by the checkpoint factor Rad9. Loss of Dot1 prevents H3-K79 methylation, leading to a checkpoint defect in the G1 phase of the cell cycle and to a reduction of checkpoint activation in mitosis, suggesting that another pathway contributes to Rad9 recruitment in M phase. We found that the replication factor Dpb11 is the keystone of this second pathway. dot1Δ dpb11-1 mutant cells are sensitive to UV or Zeocin treatment and cannot activate Rad53 if irradiated in M phase. Our data suggest that Dpb11 is held in proximity to damaged DNA through an interaction with the phosphorylated 9-1-1 complex, leading to Mec1-dependent phosphorylation of Rad9. Dpb11 is also phosphorylated after DNA damage, and this modification is lost in a nonphosphorylatable ddc1-T602A mutant. Finally, we show that, in vivo, Dpb11 cooperates with Dot1 in promoting Rad9 phosphorylation but also contributes to the full activation of Mec1 kinase. PMID:18541674

  9. ATM signaling and genomic stability in response to DNA damage

    International Nuclear Information System (INIS)

    Lavin, Martin F.; Birrell, Geoff; Chen, Philip; Kozlov, Sergei; Scott, Shaun; Gueven, Nuri

    2005-01-01

    DNA double strand breaks represent the most threatening lesion to the integrity of the genome in cells exposed to ionizing radiation and radiomimetic chemicals. Those breaks are recognized, signaled to cell cycle checkpoints and repaired by protein complexes. The product of the gene (ATM) mutated in the human genetic disorder ataxia-telangiectasia (A-T) plays a central role in the recognition and signaling of DNA damage. ATM is one of an ever growing number of proteins which when mutated compromise the stability of the genome and predispose to tumour development. Mechanisms for recognising double strand breaks in DNA, maintaining genome stability and minimizing risk of cancer are discussed

  10. Brain Stimulation Therapies

    Science.gov (United States)

    ... Magnetic Seizure Therapy Deep Brain Stimulation Additional Resources Brain Stimulation Therapies Overview Brain stimulation therapies can play ... for a shorter recovery time than ECT Deep Brain Stimulation Deep brain stimulation (DBS) was first developed ...

  11. ERK1/2 signaling plays an important role in topoisomerase II poison-induced G2/M checkpoint activation.

    Science.gov (United States)

    Kolb, Ryan H; Greer, Patrick M; Cao, Phu T; Cowan, Kenneth H; Yan, Ying

    2012-01-01

    Topo II poisons, which target topoisomerase II (topo II) to generate enzyme mediated DNA damage, have been commonly used for anti-cancer treatment. While clinical evidence demonstrate a capability of topo II poisons in inducing apoptosis in cancer cells, accumulating evidence also show that topo II poison treatment frequently results in cell cycle arrest in cancer cells, which was associated with subsequent resistance to these treatments. Results in this report indicate that treatment of MCF-7 and T47D breast cancer cells with topo II poisons resulted in an increased phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) and an subsequent induction of G2/M cell cycle arrest. Furthermore, inhibition of ERK1/2 activation using specific inhibitors markedly attenuated the topo II poison-induced G2/M arrest and diminished the topo II poison-induced activation of ATR and Chk1 kinases. Moreover, decreased expression of ATR by specific shRNA diminished topo II poison-induced G2/M arrest but had no effect on topo II poison-induced ERK1/2 activation. In contrast, inhibition of ERK1/2 signaling had little, if any, effect on topo II poison-induced ATM activation. In addition, ATM inhibition by either incubation of cells with ATM specific inhibitor or transfection of cells with ATM specific siRNA did not block topo II poison-induced G2/M arrest. Ultimately, inhibition of ERK1/2 signaling greatly enhanced topo II poison-induced apoptosis. These results implicate a critical role for ERK1/2 signaling in the activation of G2/M checkpoint response following topo II poison treatment, which protects cells from topo II poison-induced apoptosis.

  12. Different roles of the Mre11 complex in the DNA damage response in Aspergillus nidulans.

    Science.gov (United States)

    Semighini, Camile P; von Zeska Kress Fagundes, Márcia Regina; Ferreira, Joseane Cristina; Pascon, Renata Castiglioni; de Souza Goldman, Maria Helena; Goldman, Gustavo Henrique

    2003-06-01

    The Mre11-Rad50-Nbs1 protein complex has emerged as a central player in the cellular DNA damage response. Mutations in scaANBS1, which encodes the apparent homologue of human Nbs1 in Aspergillus nidulans, inhibit growth in the presence of the anti-topoisomerase I drug camptothecin. We have used the scaANBS1 cDNA as a bait in a yeast two-hybrid screening and report the identification of the A. nidulans Mre11 homologue (mreA). The inactivated mreA strain was more sensitive to several DNA damaging and oxidative stress agents. Septation in A. nidulans is dependent not only on the uvsBATR gene, but also on the mre11 complex. scaANBS1 and mreA genes are both involved in the DNA replication checkpoint whereas mreA is specifically involved in the intra-S-phase checkpoint. ScaANBS1 also participates in G2-M checkpoint control upon DNA damage caused by MMS. In addition, the scaANBS1 gene is also important for ascospore viability, whereas mreA is required for successful meiosis in A. nidulans. Consistent with this view, the Mre11 complex and the uvsCRAD51 gene are highly expressed at the mRNA level during the sexual development.

  13. Radiation damage

    CERN Document Server

    Heijne, Erik H M; CERN. Geneva

    1998-01-01

    a) Radiation damage in organic materials. This series of lectures will give an overview of radiation effects on materials and components frequently used in accelerator engineering and experiments. Basic degradation phenomena will be presented for organic materials with comprehensive damage threshold doses for commonly used rubbers, thermoplastics, thermosets and composite materials. Some indications will be given for glass, scintillators and optical fibres. b) Radiation effects in semiconductor materials and devices. The major part of the time will be devoted to treat radiation effects in semiconductor sensors and the associated electronics, in particular displacement damage, interface and single event phenomena. Evaluation methods and practical aspects will be shown. Strategies will be developed for the survival of the materials under the expected environmental conditions of the LHC machine and detectors. I will describe profound revolution in our understanding of black holes and their relation to quantum me...

  14. The Role of Rad17 in DNA Damage Checkpoint Signaling and Initiation of Apoptosis in Mammary Cells

    Science.gov (United States)

    2005-07-01

    Shen et al. 1998). Antibodies to β-catenin (Transduction lab ), GSK-3β (Santa Cruz), Smad3 I-20 (Santa Cruz), Smad3 (Zymed), phspho-Smad2 (Cell...van de Wetering, M., R. Cavallo, D. Dooijes, M. van Beest , J. van Es, J. Loureiro, A. Ypma, D. Hursh, T. Jones, A. Bejsovec, M. Peifer, M. Mortin

  15. Tort Damages

    NARCIS (Netherlands)

    L.T. Visscher (Louis)

    2008-01-01

    textabstractAbstract: In this Chapter, I provide an overview of Law and Economics literature regarding tort damages. Where necessary, attention is also spent to rules of tort liability. Both types of rules provide behavioral incentives to both injurers and victims, with respect to their level of

  16. Ultrafiltered pig leukocyte extract (UPLE, IMUNOR) potentiates hematopoiesis-stimulating effects of G-CSF in vitro and improves the outcome of treatment of hematopoietic radiation damage in mice with G-CSF

    Czech Academy of Sciences Publication Activity Database

    Vacek, Antonín; Hofer, Michal; Schneiderová, H.; Svoboda, J.

    2005-01-01

    Roč. 27, č. 4 (2005), s. 647-659 ISSN 0892-3973 R&D Projects: GA AV ČR(CZ) IBS5004009; GA AV ČR(CZ) KSK5011112 Institutional research plan: CEZ:AV0Z50040507 Keywords : ultrafiltered pig leukocyte extract * Imunor * granulocyte colony-stimulating factor Subject RIV: BO - Biophysics Impact factor: 0.557, year: 2005

  17. T cells in chronic lymphocytic leukemia display dysregulated expression of immune checkpoints and activation markers.

    Science.gov (United States)

    Palma, Marzia; Gentilcore, Giusy; Heimersson, Kia; Mozaffari, Fariba; Näsman-Glaser, Barbro; Young, Emma; Rosenquist, Richard; Hansson, Lotta; Österborg, Anders; Mellstedt, Håkan

    2017-03-01

    Chronic lymphocytic leukemia is characterized by impaired immune functions largely due to profound T-cell defects. T-cell functions also depend on co-signaling receptors, inhibitory or stimulatory, known as immune checkpoints, including cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) and programmed death-1 (PD-1). Here we analyzed the T-cell phenotype focusing on immune checkpoints and activation markers in chronic lymphocytic leukemia patients (n=80) with different clinical characteristics and compared them to healthy controls. In general, patients had higher absolute numbers of CD3 + cells and the CD8 + subset was particularly expanded in previously treated patients. Progressive patients had higher numbers of CD4 + and CD8 + cells expressing PD-1 compared to healthy controls, which was more pronounced in previously treated patients ( P =0.0003 and P =0.001, respectively). A significant increase in antigen-experienced T cells was observed in patients within both the CD4 + and CD8 + subsets, with a significantly higher PD-1 expression. Higher numbers of CD4 + and CD8 + cells with intracellular CTLA-4 were observed in patients, as well as high numbers of proliferating (Ki67 + ) and activated (CD69 + ) CD4 + and CD8 + cells, more pronounced in patients with active disease. The numbers of Th1, Th2, Th17 and regulatory T cells were substantially increased in patients compared to controls ( P leukemia T cells display increased expression of immune checkpoints, abnormal subset distribution, and a higher proportion of proliferating cells compared to healthy T cells. Disease activity and previous treatment shape the T-cell profile of chronic lymphocytic leukemia patients in different ways. Copyright© Ferrata Storti Foundation.

  18. Checkpoint Inhibition: Programmed Cell Death 1 and Programmed Cell Death 1 Ligand Inhibitors in Hodgkin Lymphoma.

    Science.gov (United States)

    Villasboas, Jose Caetano; Ansell, Stephen

    2016-01-01

    Hodgkin lymphoma (HL) is a lymphoid malignancy characterized by a reactive immune infiltrate surrounding relatively few malignant cells. In this scenario, active immune evasion seems to play a central role in allowing tumor progression. Immune checkpoint inhibitor pathways are normal mechanisms of T-cell regulation that suppress immune effector function following an antigenic challenge. Hodgkin lymphoma cells are able to escape immune surveillance by co-opting these mechanisms. The programmed cell death 1 (PD-1) pathway in particular is exploited in HL as the malignant Hodgkin and Reed-Sternberg cells express on their surface cognate ligands (PD-L1/L2) for the PD-1 receptor and thereby dampen the T-cell-mediated antitumoral response. Monoclonal antibodies that interact with and disrupt the PD-1:PD-L1/L2 axis have now been developed and tested in early-phase clinical trials in patients with advanced HL with encouraging results. The remarkable clinical activity of PD-1 inhibitors in HL highlights the importance of immune checkpoint pathways as therapeutic targets in HL. In this review, we discuss the rationale for targeting PD-1 and PD-L1 in the treatment of HL. We will evaluate the published clinical data on the different agents and highlight the safety profile of this class of agents. We discuss the available evidence on the use of biomarkers as predictors of response to checkpoint blockade and summarize the areas under active investigation in the use of PD-1/PD-L1 inhibitors for the treatment of HL.

  19. Immuno-oncologic Approaches: CAR-T Cells and Checkpoint Inhibitors.

    Science.gov (United States)

    Gay, Francesca; D'Agostino, Mattia; Giaccone, Luisa; Genuardi, Mariella; Festuccia, Moreno; Boccadoro, Mario; Bruno, Benedetto

    2017-08-01

    Advances in understanding myeloma biology have shown that disease progression is not only the consequence of intrinsic tumor changes but also of interactions between the tumor and the microenvironment in which the cancer grows. The immune system is an important component of the tumor microenvironment in myeloma, and acting on the immune system is an appealing new treatment strategy. There are 2 ways to act toward immune cells and boost antitumor immunity: (1) to increase antitumor activity (acting on T and NK cytotoxic cells), and (2) to reduce immunosuppression (acting on myeloid-derived stem cells and T regulatory cells). Checkpoint inhibitors and adoptive cell therapy (ACT) are 2 of the main actors, together with monoclonal antibodies and immunomodulatory agents, in the immune-oncologic approach. The aim of checkpoint inhibitors is to release the brakes that block the action of the immune system against the tumor. Anti-programmed death-1 (PD-1) and PD-1-Ligand, as well as anti-CTLA4 and KIR are currently under evaluation, as single agents or in combination, with the best results achieved so far with combination of anti-PD-1 and immunomodulatory agents. The aim of ACT is to create an immune effector specific against the tumor. Preliminary results on chimeric antigen receptor (CAR) T cells, first against CD19, and more recently against B-cell maturation antigen, have shown to induce durable responses in heavily pretreated patients. This review focuses on the most recent clinical results available on the use of checkpoint inhibitors and CAR-T cells in myeloma, in the context of the new immune-oncologic approach. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Requirement for PLK1 kinase activity in the maintenance of a robust spindle assembly checkpoint

    Directory of Open Access Journals (Sweden)

    Aisling O'Connor

    2016-01-01

    Full Text Available During mitotic arrest induced by microtubule targeting drugs, the weakening of the spindle assembly checkpoint (SAC allows cells to progress through the cell cycle without chromosome segregation occurring. PLK1 kinase plays a major role in mitosis and emerging evidence indicates that PLK1 is also involved in establishing the checkpoint and maintaining SAC signalling. However, mechanistically, the role of PLK1 in the SAC is not fully understood, with several recent reports indicating that it can cooperate with either one of the major checkpoint kinases, Aurora B or MPS1. In this study, we assess the role of PLK1 in SAC maintenance. We find that in nocodazole-arrested U2OS cells, PLK1 activity is continuously required for maintaining Aurora B protein localisation and activity at kinetochores. Consistent with published data we find that upon PLK1 inhibition, phosphoThr3-H3, a marker of Haspin activity, is reduced. Intriguingly, Aurora B inhibition causes PLK1 to relocalise from kinetochores into fewer and much larger foci, possibly due to incomplete recruitment of outer kinetochore proteins. Importantly, PLK1 inhibition, together with partial inhibition of Aurora B, allows efficient SAC override to occur. This phenotype is more pronounced than the phenotype observed by combining the same PLK1 inhibitors with partial MPS1 inhibition. We also find that PLK1 inhibition does not obviously cooperate with Haspin inhibition to promote SAC override. These results indicate that PLK1 is directly involved in maintaining efficient SAC signalling, possibly by cooperating in a positive feedback loop with Aurora B, and that partially redundant mechanisms exist which reinforce the SAC.

  1. Mutant p53 perturbs DNA replication checkpoint control through TopBP1 and Treslin.

    Science.gov (United States)

    Liu, Kang; Lin, Fang-Tsyr; Graves, Joshua D; Lee, Yu-Ju; Lin, Weei-Chin

    2017-05-09

    Accumulating evidence supports the gain-of-function of mutant forms of p53 (mutp53s). However, whether mutp53 directly perturbs the DNA replication checkpoint remains unclear. Previously, we have demonstrated that TopBP1 forms a complex with mutp53s and mediates their gain-of-function through NF-Y and p63/p73. Akt phosphorylates TopBP1 and induces its oligomerization, which inhibits its ATR-activating function. Here we show that various contact and conformational mutp53s bypass Akt to induce TopBP1 oligomerization and attenuate ATR checkpoint response during replication stress. The effect on ATR response caused by mutp53 can be exploited in a synthetic lethality strategy, as depletion of another ATR activator, DNA2, in mutp53-R273H-expressing cancer cells renders cells hypersensitive to cisplatin. Expression of mutp53-R273H also makes cancer cells more sensitive to DNA2 depletion or DNA2 inhibitors. In addition to ATR-activating function during replication stress, TopBP1 interacts with Treslin in a Cdk-dependent manner to initiate DNA replication during normal growth. We find that mutp53 also interferes with TopBP1 replication function. Several contact, but not conformational, mutp53s enhance the interaction between TopBP1 and Treslin and promote DNA replication despite the presence of a Cdk2 inhibitor. Together, these data uncover two distinct mechanisms by which mutp53 enhances DNA replication: ( i ) Both contact and conformational mutp53s can bind TopBP1 and attenuate the checkpoint response to replication stress, and ( ii ) during normal growth, contact (but not conformational) mutp53s can override the Cdk2 requirement to promote replication by facilitating the TopBP1/Treslin interaction.

  2. Effect of salt-inducible kinase 2 on checkpoint in response to γ-ray irradiation

    International Nuclear Information System (INIS)

    Yin Jiaojiao; Zhou Lijun; Wang Yu; Liu Xiaodan; Gu Yongqing; Zhou Pingkun

    2014-01-01

    Objective: To investigate the effect of salt-induced kinase 2 (SIK2) in the G_2/M checkpoint in response to ionizing radiation and the possible mechanism. Methods: HeLa cells were irradiated with "6"0Co γ-rays. The cell model of knockdown SIK2 expression was constrcuted by transfecting HeLa cells with a pSicoR-based lentivirus vector of expressing SIK2 shRNA by lipofectamin 2000. Western blot and flow cytometry were performed to measure the changes of SIK2 protein level and cell cycle distribution. The phosphorylated histone protein H3 on Ser 10 was used as a molecular marker of mitotic cells for detecting the function of G2/M checkpoint. Results: The expression level of SIK2 protein increased in HeLa cells after "6"0Co γ-ray irradiation. A cell model of knockdown SIK2 expression was successfully generated by transfecting the specific shRNA against SIK2. Depression of SIK2 significantly increased the cellular sensitivity at 1, 2, 4, 6 Gy post-irradiation (t = -3.445, -2.581, -3.251, -2.553, P < 0.05), and led cells to release earlier from the G_2/M boundary arrest compared to control cells at 5, 6 h post-irradiation(t = 4.341, 6.500, P < 0.05). Western blot analysis indicated that the irradiation-induced phosphorylated CHK2/T68 in SIK2 knock-down cells was earlier than that in control cells. Conclusions: salt-induced kinase 2 (SIK2) participates in the regulation of G_2/M checkpoint induced by ionizing radiation and affects cellular radiosensitivity. (authors)

  3. Monitoring of scrap loads at Gorzia border checkpoints: A thirty months experience and some suggestions

    International Nuclear Information System (INIS)

    Fabretto, M.

    1999-01-01

    The following topics are highlighted: personnel training, assistance from foreign scrap suppliers, and computerized database. It is suggested that when selecting personnel for this job, major attention should be paid to their attitudes; the personnel should be given very good training, teaching them also how to make controls manually; expert physicist' support should be available within a very short time; each monitoring checkpoint should have its local electronic database; periodical exchange and updating of information should be practised on the standardization of methods of detection, of levels of attention and alarm, of statistics among the people in charge of EU and non-EU countries. (P.A.)

  4. Strategies to overcome HBV-specific T cell exhaustion: checkpoint inhibitors and metabolic re-programming.

    Science.gov (United States)

    Fisicaro, Paola; Boni, Carolina; Barili, Valeria; Laccabue, Diletta; Ferrari, Carlo

    2018-01-29

    HBV-specific T cells play a key role in antiviral protection and failure to control HBV is associated with severely dysfunctional T cell responses. Therefore, functional T cell reconstitution represents a potential way to treat chronically infected patients. The growing understanding of the dysregulated transcriptional/epigenetic and metabolic programs underlying T cell exhaustion allows to envisage functional T cell reconstitution strategies based on the combined/sequential use of compounds able to induce decline of antigen load, checkpoint modulation, metabolic and epigenetic reprogramming with possible boosting of functionally restored responses by specific vaccines. Copyright © 2018 Elsevier B.V. All rights reserved.

  5. Response to DNA damage: why do we need to focus on protein phosphatases?

    Directory of Open Access Journals (Sweden)

    Midori eShimada

    2013-01-01

    Full Text Available Eukaryotic cells are continuously threatened by unavoidable errors during normal DNA replication or various sources of genotoxic stresses that cause DNA damage or stalled replication. To maintain genomic integrity, cells have developed a coordinated signaling network, known as the DNA damage response (DDR. Following DNA damage, sensor molecules detect the presence of DNA damage and transmit signals to downstream transducer molecules. This in turn conveys the signals to numerous effectors, which initiate a large number of specific biological responses, including transient cell cycle arrest mediated by checkpoints, DNA repair, and apoptosis. It is recently becoming clear that dephosphorylation events are involved in keeping DDR factors inactive during normal cell growth. Moreover, dephosphorylation is required to shut off checkpoint arrest following DNA damage and has been implicated in the activation of the DDR. Spatial and temporal regulation of phosphorylation events is essential for the DDR, and fine-tuning of phosphorylation is partly mediated by protein phosphatases. While the role of kinases in the DDR has been well documented, the complex roles of protein dephosphorylation have only recently begun to be investigated. Therefore, it is important to focus on the role of phosphatases and to determine how their activity is regulated upon DNA damage. In this work, we summarize current knowledge on the involvement of serine/threonine phosphatases, especially the protein phosphatase 1, protein phosphatase 2A, and protein phosphatase Mg2+/Mn2+-dependent families, in the DDR.

  6. Cross-Talk between Carbon Metabolism and the DNA Damage Response in S. cerevisiae

    Directory of Open Access Journals (Sweden)

    Kobi J. Simpson-Lavy

    2015-09-01

    Full Text Available Yeast cells with DNA damage avoid respiration, presumably because products of oxidative metabolism can be harmful to DNA. We show that DNA damage inhibits the activity of the Snf1 (AMP-activated protein kinase (AMPK, which activates expression of genes required for respiration. Glucose and DNA damage upregulate SUMOylation of Snf1, catalyzed by the SUMO E3 ligase Mms21, which inhibits SNF1 activity. The DNA damage checkpoint kinases Mec1/ATR and Tel1/ATM, as well as the nutrient-sensing protein kinase A (PKA, regulate Mms21 activity toward Snf1. Mec1 and Tel1 are required for two SNF1-regulated processes—glucose sensing and ADH2 gene expression—even without exogenous genotoxic stress. Our results imply that inhibition of Snf1 by SUMOylation is a mechanism by which cells lower their respiration in response to DNA damage. This raises the possibility that activation of DNA damage checkpoint mechanisms could contribute to aerobic fermentation (Warburg effect, a hallmark of cancer cells.

  7. DNA damage responses in human induced pluripotent stem cells and embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Olga Momcilovic

    2010-10-01

    Full Text Available Induced pluripotent stem (iPS cells have the capability to undergo self-renewal and differentiation into all somatic cell types. Since they can be produced through somatic cell reprogramming, which uses a defined set of transcription factors, iPS cells represent important sources of patient-specific cells for clinical applications. However, before these cells can be used in therapeutic designs, it is essential to understand their genetic stability.Here, we describe DNA damage responses in human iPS cells. We observe hypersensitivity to DNA damaging agents resulting in rapid induction of apoptosis after γ-irradiation. Expression of pluripotency factors does not appear to be diminished after irradiation in iPS cells. Following irradiation, iPS cells activate checkpoint signaling, evidenced by phosphorylation of ATM, NBS1, CHEK2, and TP53, localization of ATM to the double strand breaks (DSB, and localization of TP53 to the nucleus of NANOG-positive cells. We demonstrate that iPS cells temporary arrest cell cycle progression in the G(2 phase of the cell cycle, displaying a lack of the G(1/S cell cycle arrest similar to human embryonic stem (ES cells. Furthermore, both cell types remove DSB within six hours of γ-irradiation, form RAD51 foci and exhibit sister chromatid exchanges suggesting homologous recombination repair. Finally, we report elevated expression of genes involved in DNA damage signaling, checkpoint function, and repair of various types of DNA lesions in ES and iPS cells relative to their differentiated counterparts.High degrees of similarity in DNA damage responses between ES and iPS cells were found. Even though reprogramming did not alter checkpoint signaling following DNA damage, dramatic changes in cell cycle structure, including a high percentage of cells in the S phase, increased radiosensitivity and loss of DNA damage-induced G(1/S cell cycle arrest, were observed in stem cells generated by induced pluripotency.

  8. In vitro analysis of the role of replication protein A (RPA) and RPA phosphorylation in ATR-mediated checkpoint signaling.

    Science.gov (United States)

    Lindsey-Boltz, Laura A; Reardon, Joyce T; Wold, Marc S; Sancar, Aziz

    2012-10-19

    Replication protein A (RPA) plays essential roles in DNA metabolism, including replication, checkpoint, and repair. Recently, we described an in vitro system in which the phosphorylation of human Chk1 kinase by ATR (ataxia telangiectasia mutated and Rad3-related) is dependent on RPA bound to single-stranded DNA. Here, we report that phosphorylation of other ATR targets, p53 and Rad17, has the same requirements and that RPA is also phosphorylated in this system. At high p53 or Rad17 concentrations, RPA phosphorylation is inhibited and, in this system, RPA with phosphomimetic mutations cannot support ATR kinase function, whereas a non-phosphorylatable RPA mutant exhibits full activity. Phosphorylation of these ATR substrates depends on the recruitment of ATR and the substrates by RPA to the RPA-ssDNA complex. Finally, mutant RPAs lacking checkpoint function exhibit essentially normal activity in nucleotide excision repair, revealing RPA separation of function for checkpoint and excision repair.

  9. In Vitro Analysis of the Role of Replication Protein A (RPA) and RPA Phosphorylation in ATR-mediated Checkpoint Signaling*

    Science.gov (United States)

    Lindsey-Boltz, Laura A.; Reardon, Joyce T.; Wold, Marc S.; Sancar, Aziz

    2012-01-01

    Replication protein A (RPA) plays essential roles in DNA metabolism, including replication, checkpoint, and repair. Recently, we described an in vitro system in which the phosphorylation of human Chk1 kinase by ATR (ataxia telangiectasia mutated and Rad3-related) is dependent on RPA bound to single-stranded DNA. Here, we report that phosphorylation of other ATR targets, p53 and Rad17, has the same requirements and that RPA is also phosphorylated in this system. At high p53 or Rad17 concentrations, RPA phosphorylation is inhibited and, in this system, RPA with phosphomimetic mutations cannot support ATR kinase function, whereas a non-phosphorylatable RPA mutant exhibits full activity. Phosphorylation of these ATR substrates depends on the recruitment of ATR and the substrates by RPA to the RPA-ssDNA complex. Finally, mutant RPAs lacking checkpoint function exhibit essentially normal activity in nucleotide excision repair, revealing RPA separation of function for checkpoint and excision repair. PMID:22948311

  10. Targeting Tumor-Associated Macrophages as a Potential Strategy to Enhance the Response to Immune Checkpoint Inhibitors.

    Science.gov (United States)

    Cassetta, Luca; Kitamura, Takanori

    2018-01-01

    Inhibition of immune checkpoint pathways in CD8 + T cell is a promising therapeutic strategy for the treatment of solid tumors that has shown significant anti-tumor effects and is now approved by the FDA to treat patients with melanoma and lung cancer. However the response to this therapy is limited to a certain fraction of patients and tumor types, for reasons still unknown. To ensure success of this treatment, CD8 + T cells, the main target of the checkpoint inhibitors, should exert full cytotoxicity against tumor cells. However recent studies show that tumor-associated macrophages (TAM) can impede this process by different mechanisms. In this mini-review we will summarize recent studies showing the effect of TAM targeting on immune checkpoint inhibitors efficacy. We will also discuss on the limitations of the current strategies as well on the future scientific challenges for the progress of the tumor immunology field.

  11. Optical stimulation of peripheral nerves in vivo

    Science.gov (United States)

    Wells, Jonathon D.

    This dissertation documents the emergence and validation of a new clinical tool that bridges the fields of biomedical optics and neuroscience. The research herein describes an innovative method for direct neurostimulation with pulsed infrared laser light. Safety and effectiveness of this technique are first demonstrated through functional stimulation of the rat sciatic nerve in vivo. The Holmium:YAG laser (lambda = 2.12 mum) is shown to operate at an optimal wavelength for peripheral nerve stimulation with advantages over standard electrical neural stimulation; including contact-free stimulation, high spatial selectivity, and lack of a stimulation artifact. The underlying biophysical mechanism responsible for transient optical nerve stimulation appears to be a small, absorption driven thermal gradient sustained at the axonal layer of nerve. Results explicitly prove that low frequency optical stimulation can reliably stimulate without resulting in tissue thermal damage. Based on the positive results from animal studies, these optimal laser parameters were utilized to move this research into the clinic with a combined safety and efficacy study in human subjects undergoing selective dorsal rhizotomy. The clinical Holmium:YAG laser was used to effectively stimulate human dorsal spinal roots and elicit functional muscle responses recorded during surgery without evidence of nerve damage. Overall these results predict that this technology can be a valuable clinical tool in various neurosurgical applications.

  12. Grp/DChk1 is required for G(2)-M checkpoint activation in Drosophila S2 cells, whereas Dmnk/DChk2 is dispensable

    NARCIS (Netherlands)

    de Vries, HI; Uyetake, L; Lemstra, W; Brunsting, JF; Su, TT; Kampinga, HH; Sibon, OCM

    2005-01-01

    Cell-cycle checkpoints are signal-transduction pathways required to maintain genomic stability in dividing cells. Previously, it was reported that two kinases essential for checkpoint signalling, Chk1 and Chk2 are structurally conserved. In contrast to yeast, Xenopus and mammals, the Chk1- and

  13. The Geography of Deterrence: Exploring the Small Area Effects of Sobriety Checkpoints on Alcohol-Impaired Collision Rates within a City

    Science.gov (United States)

    Nunn, Samuel; Newby, William

    2011-01-01

    This article examines alcohol-impaired collision metrics around nine sobriety checkpoint locations in Indianapolis, Indiana, before and after implementation of 22 checkpoints, using a pre/post examination, a pre/post nonequivalent comparison group analysis, and an interrupted time series approach. Traffic safety officials used geographical…

  14. Nek2A destruction marks APC/C activation at the prophase-to-prometaphase transition by spindle-checkpoint-restricted Cdc20.

    Science.gov (United States)

    Boekhout, Michiel; Wolthuis, Rob

    2015-04-15

    Nek2 isoform A (Nek2A) is a presumed substrate of the anaphase-promoting complex/cyclosome containing Cdc20 (APC/C(Cdc20)). Nek2A, like cyclin A, is degraded in mitosis while the spindle checkpoint is active. Cyclin A prevents spindle checkpoint proteins from binding to Cdc20 and is recruited to the APC/C in prometaphase. We found that Nek2A and cyclin A avoid being stabilized by the spindle checkpoint in different ways. First, enhancing mitotic checkpoint complex (MCC) formation by nocodazole treatment inhibited the degradation of geminin and cyclin A, whereas Nek2A disappeared at a normal rate. Second, depleting Cdc20 effectively stabilized cyclin A but not Nek2A. Nevertheless, Nek2A destruction crucially depended on Cdc20 binding to the APC/C. Third, in contrast to cyclin A, Nek2A was recruited to the APC/C before the start of mitosis. Interestingly, the spindle checkpoint very effectively stabilized an APC/C-binding mutant of Nek2A, which required the Nek2A KEN box. Apparently, in cells, the spindle checkpoint primarily prevents Cdc20 from binding destruction motifs. Nek2A disappearance marks the prophase-to-prometaphase transition, when Cdc20, regardless of the spindle checkpoint, activates the APC/C. However, Mad2 depletion accelerated Nek2A destruction, showing that spindle checkpoint release further increases APC/C(Cdc20) catalytic activity. © 2015. Published by The Company of Biologists Ltd.

  15. RING finger and WD repeat domain 3 (RFWD3) associates with replication protein A (RPA) and facilitates RPA-mediated DNA damage response.

    Science.gov (United States)

    Liu, Shangfeng; Chu, Jessica; Yucer, Nur; Leng, Mei; Wang, Shih-Ya; Chen, Benjamin P C; Hittelman, Walter N; Wang, Yi

    2011-06-24

    DNA damage response is crucial for maintaining genomic integrity and preventing cancer by coordinating the activation of checkpoints and the repair of damaged DNA. Central to DNA damage response are the two checkpoint kinases ATM and ATR that phosphorylate a wide range of substrates. RING finger and WD repeat domain 3 (RFWD3) was initially identified as a substrate of ATM/ATR from a proteomic screen. Subsequent studies showed that RFWD3 is an E3 ubiquitin ligase that ubiquitinates p53 in vitro and positively regulates p53 levels in response to DNA damage. We report here that RFWD3 associates with replication protein A (RPA), a single-stranded DNA-binding protein that plays essential roles in DNA replication, recombination, and repair. Binding of RPA to single-stranded DNA (ssDNA), which is generated by DNA damage and repair, is essential for the recruitment of DNA repair factors to damaged sites and the activation of checkpoint signaling. We show that RFWD3 is physically associated with RPA and rapidly localizes to sites of DNA damage in a RPA-dependent manner. In vitro experiments suggest that the C terminus of RFWD3, which encompass the coiled-coil domain and the WD40 domain, is necessary for binding to RPA. Furthermore, DNA damage-induced phosphorylation of RPA and RFWD3 is dependent upon each other. Consequently, loss of RFWD3 results in the persistent foci of DNA damage marker γH2AX and the repair protein Rad51 in damaged cells. These findings suggest that RFWD3 is recruited to sites of DNA damage and facilitates RPA-mediated DNA damage signaling and repair.

  16. Moderate variations in CDC25B protein levels modulate the response to DNA damaging agents

    International Nuclear Information System (INIS)

    Aressy, B.; Bugler, B.; Valette, A.; Ducommun, B.; Biard, D.

    2008-01-01

    CDC25B, one of the three members of the CDC25 dual-specificity phosphatase family, plays a critical role in the control of the cell cycle and in the checkpoint response to DNA damage. CDC25B is responsible for the initial dephosphorylation and activation of the cyclin-dependent kinases, thus initiating the train of events leading to entry into mitosis. The critical role played by CDC25B is illustrated by the fact that it is specifically required for checkpoint recovery and that unscheduled accumulation of CDC25B is responsible for illegitimate entry into mitosis. Here, we report that in p53 colon carcinoma cells, a moderate increase in the CDC25B level is sufficient to impair the DNA damage checkpoint, to increase spontaneous mutagenesis, and to sensitize cells to ionising radiation and genotoxic agents. Using a tumour cell spheroid assay as an alternative to animal studies, we demonstrate that the level of CDC25B expression modulates growth inhibition and apoptotic death. Since CDC25B overexpression has been observed in a significant number of human cancers, including colon carcinoma, and is often associated with high grade tumours and poor prognosis, our work suggests that the expression level of CDC25B might be a potential key parameter of the cellular response to cancer therapy. (authors)

  17. Structural basis of a novel PD-L1 nanobody for immune checkpoint blockade.

    Science.gov (United States)

    Zhang, Fei; Wei, Hudie; Wang, Xiaoxiao; Bai, Yu; Wang, Pilin; Wu, Jiawei; Jiang, Xiaoyong; Wang, Yugang; Cai, Haiyan; Xu, Ting; Zhou, Aiwu

    2017-01-01

    The use of antibodies to target immune checkpoints, particularly PD-1/PD-L1, has made a profound impact in the field of cancer immunotherapy. Here, we identified KN035, an anti-PD-L1 nanobody that can strongly induce T-cell responses and inhibit tumor growth. The crystal structures of KN035 complexed with PD-L1 and free PD-L1, solved here at 1.7 and 2.7 Å resolution, respectively, show that KN035 competes with PD-1 (programmed death protein 1) for the same flat surface on PD-L1, mainly through a single surface loop of 21 amino acids. This loop forms two short helices and develops key hydrophobic and ionic interactions with PD-L1 residues, such as Ile54, Tyr56 and Arg113, which are also involved in PD-1 binding. The detailed mutagenesis study identified the hotspot residues of the PD-L1 surface and provides an explanation for the stronger (~1 000-fold) binding of KN035 to PD-L1 than PD-1 and its lack of binding to PD-L2. Overall, this study reveals how a single immunoglobulin-variable scaffold of KN035 or PD-1 can bind to a flat protein surface through either a single surface loop or beta-sheet strands; and provides a basis for designing new immune checkpoint blockers and generating bi-specific antibodies for combination therapy.

  18. Nucleotide Selectivity at a Preinsertion Checkpoint of T7 RNA Polymerase Transcription Elongation.

    Science.gov (United States)

    E, Chao; Duan, Baogen; Yu, Jin

    2017-04-20

    Nucleotide selection is crucial for transcription fidelity control, in particular, for viral T7 RNA polymerase (RNAP) lack of proofreading activity. It has been recognized that multiple kinetic checkpoints exist prior to full nucleotide incorporation. In this work, we implemented intensive atomistic molecular dynamics (MD) simulations to quantify how strong the nucleotide selection is at the initial checkpoint of an elongation cycle of T7 RNAP. The incoming nucleotides bind into a preinsertion site where a critical tyrosine residue locates nearby to assist the nucleotide selection. We calculated the relative binding free energy between a noncognate nucleotide and a cognate one at a preinsertion configuration via alchemical simulations, showing that a small selection free energy or the binding free energy difference (∼3 k B T) exists between the two nucleotides. Indeed, another preinsertion configuration favored by the noncognate nucleotides was identified, which appears to be off path for further nucleotide insertion and additionally assists the nucleotide selection. By chemical master equation (CME) approach, we show that the small selection free energy at the preinsertion site along with the off-path noncognate nucleotide filtering can help substantially to reduce the error rate and to maintain the elongation rate high in the T7 RNAP transcription.

  19. Checkpoint-dependent RNR induction promotes fork restart after replicative stress.

    Science.gov (United States)

    Morafraile, Esther C; Diffley, John F X; Tercero, José Antonio; Segurado, Mónica

    2015-01-20

    The checkpoint kinase Rad53 is crucial to regulate DNA replication in the presence of replicative stress. Under conditions that interfere with the progression of replication forks, Rad53 prevents Exo1-dependent fork degradation. However, although EXO1 deletion avoids fork degradation in rad53 mutants, it does not suppress their sensitivity to the ribonucleotide reductase (RNR) inhibitor hydroxyurea (HU). In this case, the inability to restart stalled forks is likely to account for the lethality of rad53 mutant cells after replication blocks. Here we show that Rad53 regulates replication restart through the checkpoint-dependent transcriptional response, and more specifically, through RNR induction. Thus, in addition to preventing fork degradation, Rad53 prevents cell death in the presence of HU by regulating RNR-expression and localization. When RNR is induced in the absence of Exo1 and RNR negative regulators, cell viability of rad53 mutants treated with HU is increased and the ability of replication forks to restart after replicative stress is restored.

  20. Immune checkpoint inhibitors for non-small-cell lung cancer: does that represent a 'new frontier'?

    Science.gov (United States)

    Pilotto, Sara; Kinspergher, Stefania; Peretti, Umberto; Calio, Anna; Carbognin, Luisa; Ferrara, Roberto; Brunelli, Matteo; Chilosi, Marco; Tortora, Giampaolo; Bria, Emilio

    2015-01-01

    Advances in the interpretation and understanding of cancer behaviour, particularly of its ability to evade the host immunosurveillance, deregulating the balance between inhibitory and stimulatory factors, led to the development of an innovative category of immunotherapeutic agents, currently under investigation. Although the disappointing data deriving from the employment of vaccines in non-small cell lung cancer (NSCLC), more promising results have been obtained in the early phase trials with immune checkpoint inhibitors, such as cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), programmed cell death protein-1 (PD-1) and programmed death-ligand 1 (PD-L1) inhibitors. This review delineates the main features of the available immunotherapeutic agents, focusing the discussion on immune checkpoint inhibitors, those that have already demonstrated a relevant clinical activity (such as Ipilimumab and Nivolumab) and those molecules still in early development phase. Moreover, we underline the possible emerging issues deriving from the progressive diffusion of Immuno-Oncology into the standard clinical practice. The careful and accurate identification and management of immune-related toxicities, the validation of more reliable immune response criteria and the increasing research of potential predictive biomarkers are key points of discussion. The perspective is that immunotherapy might represent an effective 'magic bullet', able to change the treatment paradigm of NSCLC, particularly of those subgroups featured by a heavily mutant cancer (squamous histology and smokers), where the immunologic agents contribute in cancer development and progression seems to be strong and, concurrently, the efficacy of standard therapies particularly limited.

  1. Preferential radiosensitization of G1 checkpoint--deficient cells by methylxanthines

    International Nuclear Information System (INIS)

    Russell, Kenneth J.; Wiens, Linda W.; Demers, G. William; Galloway, Denise A.; Le, Tiep; Rice, Glenn C.; Bianco, James A.; Singer, Jack W.; Groudine, Mark

    1996-01-01

    Purpose: To develop a checkpoint-based strategy for preferential radiosensitization of human tumors with deficient and/or mutant p53. Methods and Materials: A549 human lung adenocarcinoma cell lines differing in their expression of the p53 tumor suppressor gene were produced by transduction with the E6 oncogene from human papilloma virus type 16. The cells expressing E6 (E6+) lack a G1 arrest in response to ionizing radiation, are deficient in p53 and p21 expression, and exhibit a fivefold greater clonogenic survival following 10 Gy radiation. Results: Postirradiation incubation with millimolar concentrations of the methylxanthine pentoxifylline (PTX) results in preferential radiosensitization of the E6+ cells compared to the LXSN+ vector transduced controls. There is a threefold sensitization of the LXSN+ cells and a 15-fold sensitization of the E6+ cells, which results in equal clonogenic survival of the two lines. Flow cytometry reveals PTX abrogation of the radiation induced G2 arrest for both cell lines. PTX also prolongs G1 transit for both cell lines. Preliminary results are presented using a novel methylxanthine, lisofylline (LSF), which has similar cell cycle effects on G1 and G2 and achieves differential radiosensitization at micromolar concentrations that are sustainable in humans. Conclusions: This checkpoint-based strategy is a promising approach for achieving preferential radiosensitization of p53- tumors relative to p53+ normal tissues

  2. Hepatic neuregulin 4 signaling defines an endocrine checkpoint for steatosis-to-NASH progression

    Science.gov (United States)

    Guo, Liang; Chen, Zhimin; Xia, Houjun; Li, Siming; Zhang, Yanqiao; Kobberup, Sune; Zou, Weiping; Lin, Jiandie D.

    2017-01-01

    Nonalcoholic steatohepatitis (NASH) is characterized by progressive liver injury, inflammation, and fibrosis; however, the mechanisms that govern the transition from hepatic steatosis, which is relatively benign, to NASH remain poorly defined. Neuregulin 4 (Nrg4) is an adipose tissue–enriched endocrine factor that elicits beneficial metabolic effects in obesity. Here, we show that Nrg4 is a key component of an endocrine checkpoint that preserves hepatocyte health and counters diet-induced NASH in mice. Nrg4 deficiency accelerated liver injury, fibrosis, inflammation, and cell death in a mouse model of NASH. In contrast, transgenic expression of Nrg4 in adipose tissue alleviated diet-induced NASH. Nrg4 attenuated hepatocyte death in a cell-autonomous manner by blocking ubiquitination and proteasomal degradation of c-FLIPL, a negative regulator of cell death. Adeno-associated virus–mediated (AAV-mediated) rescue of hepatic c-FLIPL expression in Nrg4-deficent mice functionally restored the brake for steatosis to NASH transition. Thus, hepatic Nrg4 signaling serves as an endocrine checkpoint for steatosis-to-NASH progression by activating a cytoprotective pathway to counter stress-induced liver injury. PMID:29106384

  3. Molecular basis of APC/C regulation by the spindle assembly checkpoint

    Science.gov (United States)

    Zhang, Ziguo; Yang, Jing; Maslen, Sarah; Skehel, Mark; Barford, David

    2016-01-01

    In the dividing eukaryotic cell the spindle assembly checkpoint (SAC) ensures each daughter cell inherits an identical set of chromosomes. The SAC coordinates the correct attachment of sister chromatid kinetochores to the mitotic spindle with activation of the anaphase-promoting complex/cyclosome (APC/C), the E3 ubiquitin ligase that initiates chromosome separation. In response to unattached kinetochores, the SAC generates the mitotic checkpoint complex (MCC), a multimeric assembly that inhibits the APC/C, delaying chromosome segregation. Here, using cryo-electron microscopy we determined the near-atomic resolution structure of an APC/C-MCC complex (APC/CMCC). We reveal how degron-like sequences of the MCC subunit BubR1 block degron recognition sites on Cdc20, the APC/C coactivator subunit (Cdc20APC/C) responsible for substrate interactions. BubR1 also obstructs binding of UbcH10 (APC/C’s initiating E2) to repress APC/C ubiquitination activity. Conformational variability of the complex allows for UbcH10 association, and we show from a structure of APC/CMCC in complex with UbcH10 how the Cdc20 subunit intrinsic to the MCC (Cdc20MCC) is ubiquitinated, a process that results in APC/C reactivation when the SAC is silenced. PMID:27509861

  4. The impact of aviation checkpoint queues on optimizing security screening effectiveness

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Adrian J., E-mail: ajlee@citeri.or [Central Illinois Technology and Education Research Institute, 2312 Connie Drive, Springfield, IL 62704-8722 (United States); Jacobson, Sheldon H., E-mail: shj@illinois.ed [Department of Computer Science, University of Illinois at Urbana-Champaign, 201 N. Goodwin Ave. M/C 258, Urbana, IL 61801-2302 (United States)

    2011-08-15

    Passenger screening at aviation security checkpoints is a critical component in protecting airports and aircraft from terrorist threats. Recent developments in screening device technology have increased the ability to detect these threats; however, the average amount of time it takes to screen a passenger still remains a concern. This paper models the queueing process for a multi-level airport checkpoint security system, where multiple security classes are formed through subsets of specialized screening devices. An optimal static assignment policy is obtained which minimizes the steady-state expected amount of time a passenger spends in the security system. Then, an optimal dynamic assignment policy is obtained through a transient analysis that balances the expected number of true alarms with the expected amount of time a passenger spends in the security system. Performance of a two-class system is compared to that of a selective security system containing primary and secondary levels of screening. The key contribution is that the resulting optimal assignment policies increase security and passenger throughput by efficiently and effectively utilizing available screening resources.

  5. The impact of aviation checkpoint queues on optimizing security screening effectiveness

    International Nuclear Information System (INIS)

    Lee, Adrian J.; Jacobson, Sheldon H.

    2011-01-01

    Passenger screening at aviation security checkpoints is a critical component in protecting airports and aircraft from terrorist threats. Recent developments in screening device technology have increased the ability to detect these threats; however, the average amount of time it takes to screen a passenger still remains a concern. This paper models the queueing process for a multi-level airport checkpoint security system, where multiple security classes are formed through subsets of specialized screening devices. An optimal static assignment policy is obtained which minimizes the steady-state expected amount of time a passenger spends in the security system. Then, an optimal dynamic assignment policy is obtained through a transient analysis that balances the expected number of true alarms with the expected amount of time a passenger spends in the security system. Performance of a two-class system is compared to that of a selective security system containing primary and secondary levels of screening. The key contribution is that the resulting optimal assignment policies increase security and passenger throughput by efficiently and effectively utilizing available screening resources.

  6. Patterns of DNA damage response in intracranial germ cell tumors versus glioblastomas reflect cell of origin rather than brain environment

    DEFF Research Database (Denmark)

    Bartkova, Jirina; Hoei-Hansen, Christina E; Krizova, Katerina

    2014-01-01

    The DNA damage response (DDR) machinery becomes commonly activated in response to oncogenes and during early stages of development of solid malignancies, with an exception of testicular germ cell tumors (TGCTs). The active DDR signaling evokes cell death or senescence but this anti-tumor barrier ...... checkpoints in intracranial tumorigenesis, with implications for the differential biological responses of diverse tumor types to endogenous stress as well as to genotoxic treatments such as ionizing radiation or chemotherapy....

  7. Repair response for DNA double-strand damage through ubiquitylation of chromatin

    International Nuclear Information System (INIS)

    Nakada, Shinichiro

    2011-01-01

    The chromatin modulation (remodeling) via lysine63 (K63)-linked ubiquitin (U) has been found important in the repair response for DNA double-strand damage, and the sequential signaling events at the damage site are explained. As the first step of the repair, MRN (MRE11, RAD50 and nibrin) complex recognizes the damage site and binds to it followed by many linked reactions by recruited and activated enzymes of various protein kinases and phosphatases, which resulting in the enhanced early signaling. As well, gamma-H2AX (phosphorylated histone H2AX) is yielded by the process, to which phosphorylated MDC1 (mediator of DNA-damage checkpoint 1) binds to produce their complex. Then further binding of RNF8-HERC2-UBC13 (ring finger protein 8, hect domain and RCC1 (CHC1)-like domain, and U conjugating enzyme E2N, respectively) occurs for starting the cumulative ubiquitylation of H2AX via K63 as the middle phase response. Signaling in the late phase occurs on the U chain formed at the damage site by binding of RAP (receptor-associated protein) 80 and other recruited 5 proteins like BRCA1 (breast cancer 1, early onset) to repair DNA by the homologous recombination after 53BP1 (tumor protein p53 binding protein) binding followed by methylation of histone H4. In a case of human compound heterozygous RNF168 defect, RIDDLE syndrome (radiosensitivity, immunodeficiency, dysmorphic features and learning difficulties), cells have no and slight abnormality of G2/M and intra-S checkpoint, respectively. Another defecting case with homozygous nonsense mutation has high radiosensitivity, intra-S checkpoint abnormality and others. Abnormality of immuno-globulins observed in both cases is similar to that in the RNF8-knockout mouse. Many tasks in chromatin ubiquitylation in the repair are still remained to be solved for protection and treatment of related diseases. (T.T.)

  8. ATM directs DNA damage responses and proteostasis via genetically separable pathways.

    Science.gov (United States)

    Lee, Ji-Hoon; Mand, Michael R; Kao, Chung-Hsuan; Zhou, Yi; Ryu, Seung W; Richards, Alicia L; Coon, Joshua J; Paull, Tanya T

    2018-01-09

    The protein kinase ATM is a master regulator of the DNA damage response but also responds directly to oxidative stress. Loss of ATM causes ataxia telangiectasia, a neurodegenerative disorder with pleiotropic symptoms that include cerebellar dysfunction, cancer, diabetes, and premature aging. We genetically separated the activation of ATM by DNA damage from that by oxidative stress using separation-of-function mutations. We found that deficient activation of ATM by the Mre11-Rad50-Nbs1 complex and DNA double-strand breaks resulted in loss of cell viability, checkpoint activation, and DNA end resection in response to DNA damage. In contrast, loss of oxidative activation of ATM had minimal effects on DNA damage-related outcomes but blocked ATM-mediated initiation of checkpoint responses after oxidative stress and resulted in deficiencies in mitochondrial function and autophagy. In addition, expression of a variant ATM incapable of activation by oxidative stress resulted in widespread protein aggregation. These results indicate a direct relationship between the mechanism of ATM activation and its effects on cellular metabolism and DNA damage responses in human cells and implicate ATM in the control of protein homeostasis. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  9. In vivo charge injection limits increased after 'unsafe' stimulation

    DEFF Research Database (Denmark)

    Meijs, Suzan; Sørensen, Søren; Rechendorff, Kristian

    2015-01-01

    was not observed with any of the stimulation protocols and no tissue damage was observed for the 20 mA – 200 Hz stimulation group. This indicates that the ‘safe potential window’ may not be applicable in vivo, as no damage was done stimulating with 20 mA at 200 Hz, while damage was done using the same current......The effect of unsafe stimulation on charge injection limits (Qinj) and pulsing capacitance (Cpulse) was investigated. Four stimulation protocols were applied: 20 mA – 200 and 400 Hz, 50 mA – 200 and 400 Hz. Increasing Qinj and Cpulse were observed for all stimulation protocols. Corrosion...

  10. Irradiation damage

    Energy Technology Data Exchange (ETDEWEB)

    Howe, L.M

    2000-07-01

    There is considerable interest in irradiation effects in intermetallic compounds from both the applied and fundamental aspects. Initially, this interest was associated mainly with nuclear reactor programs but it now extends to the fields of ion-beam modification of metals, behaviour of amorphous materials, ion-beam processing of electronic materials, and ion-beam simulations of various kinds. The field of irradiation damage in intermetallic compounds is rapidly expanding, and no attempt will be made in this chapter to cover all of the various aspects. Instead, attention will be focused on some specific areas and, hopefully, through these, some insight will be given into the physical processes involved, the present state of our knowledge, and the challenge of obtaining more comprehensive understanding in the future. The specific areas that will be covered are: point defects in intermetallic compounds; irradiation-enhanced ordering and irradiation-induced disordering of ordered alloys; irradiation-induced amorphization.

  11. Irradiation damage

    International Nuclear Information System (INIS)

    Howe, L.M.

    2000-01-01

    There is considerable interest in irradiation effects in intermetallic compounds from both the applied and fundamental aspects. Initially, this interest was associated mainly with nuclear reactor programs but it now extends to the fields of ion-beam modification of metals, behaviour of amorphous materials, ion-beam processing of electronic materials, and ion-beam simulations of various kinds. The field of irradiation damage in intermetallic compounds is rapidly expanding, and no attempt will be made in this chapter to cover all of the various aspects. Instead, attention will be focused on some specific areas and, hopefully, through these, some insight will be given into the physical processes involved, the present state of our knowledge, and the challenge of obtaining more comprehensive understanding in the future. The specific areas that will be covered are: point defects in intermetallic compounds; irradiation-enhanced ordering and irradiation-induced disordering of ordered alloys; irradiation-induced amorphization

  12. Checkpoint inhibitors and radiation treatment in Hodgkin's lymphoma. New study concepts of the German Hodgkin Study Group

    Energy Technology Data Exchange (ETDEWEB)

    Baues, C.; Semrau, R.; Marnitz, S. [University of Cologne, Medical Faculty, Department of Radiooncology, Cologne (Germany); University of Cologne, German Hodgkin Study Group (GHSG), Cologne (Germany); Gaipl, U.S. [University Hospital Erlangen, Department of Radiooncology, Erlangen (Germany); Broeckelmann, P.J.; Engert, A. [University of Cologne, German Hodgkin Study Group (GHSG), Cologne (Germany); University of Cologne, Medical Faculty, Department of Internal Medicine I, Cologne (Germany); Rosenbrock, J. [University of Cologne, Medical Faculty, Department of Radiooncology, Cologne (Germany)

    2017-02-15

    Patients with classical Hodgkin's lymphoma (cHL) have a good prognosis even in advanced stages. However, combined chemo- and radiotherapy, as the standard of care, is also associated with treatment-related toxicities such as organ damage, secondary neoplasias, infertility, or fatigue and long-term fatigue. Many patients suffer from this burden although their cHL was cured. Therefore, the efficacy of immune checkpoint inhibitors like anti-PD1/PD-L1 antibodies in the treatment of solid cancers and also in HL offers new options. A remarkable and durable response rate with a favorable toxicity profile was observed in heavily pretreated cHL patients. Planning to perform prospective randomized clinical trials in the content of radio-immune treatment in patients with Hodgkin's lymphoma (HL), we transferred the results of preliminary clinical studies and basic research in clinical relevant study concepts. Based on these promising early phase trial data, the German Hodgkin Study Group (GHSG) will investigate innovative treatment regimens in upcoming phase II trials. The therapeutic efficacy and potential synergies of anti-PD1 antibodies in combination with chemo- or radiotherapy will be investigated in various settings of HL. (orig.) [German] Patienten mit einem klassischen Hodgkin-Lymphom (cHL) haben ueber alle Stadien hinweg eine gute Prognose. Allerdings treten unter der kombinierten Therapie mit Chemotherapie und Bestrahlung therapieabhaengige Toxizitaeten wie z. B. Organschaeden, Sekundaertumoren, Fatigue oder Langzeit-Fatigue auf. Viele Patienten leiden trotz einer Heilung an diesen Symptomen. Daher bietet die nachgewiesene Wirksamkeit der Anti-PD1/PD-L1-Antikoerper bei soliden Tumoren, aber auch beim HL neue Behandlungsoptionen. Bei intensiv vorbehandelten Patienten mit rezidiviertem cHL wurde bei guter Vertraeglichkeit eine hohe Ansprechrate mit z. T. langanhaltenden Remissionen beobachtet. Im Rahmen der Planung prospektiver randomisierter Studien im

  13. Centromere replication timing determines different forms of genomic instability in Saccharomyces cerevisiae checkpoint mutants during replication stress.

    Science.gov (United States)

    Feng, Wenyi; Bachant, Jeff; Collingwood, David; Raghuraman, M K; Brewer, Bonita J

    2009-12-01

    Yeast replication checkpoint mutants lose viability following transient exposure to hydroxyurea, a replication-impeding drug. In an effort to understand the basis for this lethality, we discovered that different events are responsible for inviability in checkpoint-deficient cells harboring mutations in the mec1 and rad53 genes. By monitoring genomewide replication dynamics of cells exposed to hydroxyurea, we show that cells with a checkpoint deficient allele of RAD53, rad53K227A, fail to duplicate centromeres. Following removal of the drug, however, rad53K227A cells recover substantial DNA replication, including replication through centromeres. Despite this recovery, the rad53K227A mutant fails to achieve biorientation of sister centromeres during recovery from hydroxyurea, leading to secondary activation of the spindle assembly checkpoint (SAC), aneuploidy, and lethal chromosome segregation errors. We demonstrate that cell lethality from this segregation defect could be partially remedied by reinforcing bipolar attachment. In contrast, cells with the mec1-1 sml1-1 mutations suffer from severely impaired replication resumption upon removal of hydroxyurea. mec1-1 sml1-1 cells can, however, duplicate at least some of their centromeres and achieve bipolar attachment, leading to abortive segregation and fragmentation of incompletely replicated chromosomes. Our results highlight the importance of replicating yeast centromeres early and reveal different mechanisms of cell death due to differences in replication fork progression.

  14. Mad2 binding to Mad1 and Cdc20, rather than oligomerization, is required for the spindle checkpoint

    DEFF Research Database (Denmark)

    Sironi, L; Melixetian, M; Faretta, M

    2001-01-01

    Mad2 is a key component of the spindle checkpoint, a device that controls the fidelity of chromosome segregation in mitosis. The ability of Mad2 to form oligomers in vitro has been correlated with its ability to block the cell cycle upon injection into Xenopus embryos. Here we show that Mad2 forms...

  15. Structural Biology of the Immune Checkpoint Receptor PD-1 and Its Ligands PD-L1/PD-L2

    NARCIS (Netherlands)

    Zak, Krzysztof M.; Grudnik, Przemyslaw; Magiera, Katarzyna; Dömling, Alexander; Dubin, Grzegorz; Holak, Tad A.

    2017-01-01

    Cancer cells can avoid and suppress immune responses through activation of inhibitory immune checkpoint proteins, such as PD-1, PD-L1, and CTLA-4. Blocking the activities of these proteins with monoclonal antibodies, and thus restoring T cell function, has delivered breakthrough therapies against

  16. ATM/Wip1 activities at chromatin control Plk1 re-activation to determine G2 checkpoint duration

    Czech Academy of Sciences Publication Activity Database

    Jaiswal, H.; Benada, Jan; Müllers, E.; Akopyan, K.; Burdová, Kamila; Koolmeister, T.; Helleday, T.; Medema, R.H.; Macůrek, Libor; Lindqvist, A.

    2017-01-01

    Roč. 36, č. 14 (2017), s. 2161-2176 ISSN 0261-4189 R&D Projects: GA ČR GA13-18392S Institutional support: RVO:68378050 Keywords : ATM * ATR * checkpoint recovery * G2 * Pik1 Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Cell biology Impact factor: 9.792, year: 2016

  17. Design Optimization of Time- and Cost-Constrained Fault-Tolerant Embedded Systems with Checkpointing and Replication

    DEFF Research Database (Denmark)

    Pop, Paul; Izosimov, Viacheslav; Eles, Petru

    2009-01-01

    We present an approach to the synthesis of fault-tolerant hard real-time systems for safety-critical applications. We use checkpointing with rollback recovery and active replication for tolerating transient faults. Processes and communications are statically scheduled. Our synthesis approach deci...

  18. ProAtlantic - The Atlantic Checkpoint - Data Availability and Adequacy in the Atlantic Basin

    Science.gov (United States)

    McGrath, F.

    2017-12-01

    DG MAREs Atlantic Checkpoint is a basin scale wide monitoring system assessment activity based upon targeted end-user applications. It is designed to be a benchmark for the assessment of hydrographic, geological, habitat, climate and fisheries data existence and availability in the Atlantic basin. DG MAREs Atlantic Checkpoint service will be delivered by the ProAtlantic project. The objective of this project is to investigate, through appropriate methodologies in the framework of 11 key marine challenges, how current international and national data providers - e.g. EMODNet, Copernicus - meet the requirements of the stakeholders and deliver fit for purpose data. By so doing, the main thematic and geographic gaps will be readily identified in the Atlantic basin for future consideration by DG MARE. For each challenge, specific web products in the form of maps, metadata, spreadsheets and reports will be delivered. These products are not an end by themselves but rather a means of showing whether data were available, let alone accessible. For example, the Fisheries Impact Challenge outputs include data grids (VMS/Seabed) and data adequacy reports. Production of gridded data layers in order to show the extent of fisheries impact on the seafloor involved the identification, acquisition and collation of data sources for the required data types (VMS/Seabed/Habitats Data) in the Atlantic basin. The resulting spatial coverage of these grids indicates the relatively low level of data availability and adequacy across the Atlantic basin. Aside from the data delivered by programmes such as EMODNet and Copernicus, there are a lot of initiatives by regional bodies such as OSPAR and ICES that consist of assembling and disseminating data to address specific issues. Several international projects have delivered research, data collection, and networking around several of the Atlantic Checkpoint challenge topics, namely MPAs, renewable energy assessment, seabed mapping, oil spill

  19. Imaging of hepatic toxicity of systemic therapy in a tertiary cancer centre: chemotherapy, haematopoietic stem cell transplantation, molecular targeted therapies, and immune checkpoint inhibitors.

    Science.gov (United States)

    Alessandrino, F; Tirumani, S H; Krajewski, K M; Shinagare, A B; Jagannathan, J P; Ramaiya, N H; Di Salvo, D N

    2017-07-01

    The purpose of this review is to familiarise radiologists with the spectrum of hepatic toxicity seen in the oncology setting, in view of the different systemic therapies used in cancer patients. Drug-induced liver injury can manifest in various forms, and anti-neoplastic agents are associated with different types of hepatotoxicity. Although chemotherapy-induced liver injury can present as hepatitis, steatosis, sinusoidal obstruction syndrome, and chronic parenchymal damages, molecular targeted therapy-associated liver toxicity ranges from mild liver function test elevation to fulminant life-threatening acute liver failure. The recent arrival of immune checkpoint inhibitors in oncology has introduced a new range of immune-related adverse events, with differing mechanisms of liver toxicity and varied imaging presentation of liver injury. High-dose chemotherapy regimens for haematopoietic stem cell transplantation are associated with sinusoidal obstruction syndrome. Management of hepatic toxicity depends on the clinical scenario, the drug in use, and the severity of the findings. In this article, we will (1) present the most common types of oncological drugs associated with hepatic toxicity and associated liver injuries; (2) illustrate imaging findings of hepatic toxicities and the possible differential diagnosis; and (3) provide a guide for management of these conditions. Copyright © 2017 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.

  20. Phyllanthus emblica Fruit Extract Activates Spindle Assembly Checkpoint, Prevents Mitotic Aberrations and Genomic Instability in Human Colon Epithelial NCM460 Cells

    Directory of Open Access Journals (Sweden)

    Xihan Guo

    2016-09-01

    Full Text Available The fruit of Phyllanthus emblica Linn. (PE has been widely consumed as a functional food and folk medicine in Southeast Asia due to its remarkable nutritional and pharmacological effects. Previous research showed PE delays mitotic progress and increases genomic instability (GIN in human colorectal cancer cells. This study aimed to investigate the similar effects of PE by the biomarkers related to spindle assembly checkpoint (SAC, mitotic aberrations and GIN in human NCM460 normal colon epithelial cells. Cells were treated with PE and harvested differently according to the biomarkers observed. Frequencies of micronuclei (MN, nucleoplasmic bridge (NPB and nuclear bud (NB in cytokinesis-block micronucleus assay were used as indicators of GIN. Mitotic aberrations were assessed by the biomarkers of chromosome misalignment, multipolar division, chromosome lagging and chromatin bridge. SAC activity was determined by anaphase-to- metaphase ratio (AMR and the expression of core SAC gene budding uninhibited by benzimidazoles related 1 (BubR1. Compared with the control, PE-treated cells showed (1 decreased incidences of MN, NPB and NB (p < 0.01; (2 decreased frequencies of all mitotic aberration biomarkers (p < 0.01; and (3 decreased AMR (p < 0.01 and increased BubR1 expression (p < 0.001. The results revealed PE has the potential to protect human normal colon epithelial cells from mitotic and genomic damages partially by enhancing the function of SAC.

  1. Targeting cytokine signaling checkpoint CIS activates NK cells to protect from tumor initiation and metastasis

    Science.gov (United States)

    Putz, Eva M.; Guillerey, Camille; Kos, Kevin; Stannard, Kimberley; Miles, Kim; Delconte, Rebecca B.; Nicholson, Sandra E.; Huntington, Nicholas D.; Smyth, Mark J.

    2017-01-01

    ABSTRACT The cytokine-induced SH2-containing protein CIS belongs to the suppressor of cytokine signaling (SOCS) protein family. Here, we show the critical role of CIS in suppressing natural killer (NK) cell control of tumor initiation and metastasis. Cish-deficient mice were highly resistant to methylcholanthrene-induced sarcoma formation and protected from lung metastasis of B16F10 melanoma and RM-1 prostate carcinoma cells. In contrast, the growth of primary subcutaneous tumors, including those expressing the foreign antigen OVA, was unchanged in Cish-deficient mice. The combination of Cish deficiency and relevant targeted and immuno-therapies such as combined BRAF and MEK inhibitors, immune checkpoint blockade antibodies, IL-2 and type I interferon revealed further improved control of metastasis. The data clearly indicate that targeting CIS promotes NK cell antitumor functions and CIS holds great promise as a novel target in NK cell immunotherapy. PMID:28344878

  2. Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade

    Science.gov (United States)

    McGranahan, Nicholas; Furness, Andrew J. S.; Rosenthal, Rachel; Ramskov, Sofie; Lyngaa, Rikke; Saini, Sunil Kumar; Jamal-Hanjani, Mariam; Wilson, Gareth A.; Birkbak, Nicolai J.; Hiley, Crispin T.; Watkins, Thomas B. K.; Shafi, Seema; Murugaesu, Nirupa; Mitter, Richard; Akarca, Ayse U.; Linares, Joseph; Marafioti, Teresa; Henry, Jake Y.; Van Allen, Eliezer M.; Miao, Diana; Schilling, Bastian; Schadendorf, Dirk; Garraway, Levi A.; Makarov, Vladimir; Rizvi, Naiyer A.; Snyder, Alexandra; Hellmann, Matthew D.; Merghoub, Taha; Wolchok, Jedd D.; Shukla, Sachet A.; Wu, Catherine J.; Peggs, Karl S.; Chan, Timothy A.; Hadrup, Sine R.; Quezada, Sergio A.; Swanton, Charles

    2016-01-01

    As tumors grow, they acquire mutations, some of which create neoantigens that influence the response of patients to immune checkpoint inhibitors. We explored the impact of neoantigen intratumor heterogeneity (ITH) on antitumor immunity. Through integrated analysis of ITH and neoantigen burden, we demonstrate a relationship between clonal neoantigen burden and overall survival in primary lung adenocarcinomas. CD8+ tumor-infiltrating lymphocytes reactive to clonal neoantigens were identified in early-stage non–small cell lung cancer and expressed high levels of PD-1. Sensitivity to PD-1 and CTLA-4 blockade in patients with advanced NSCLC and melanoma was enhanced in tumors enriched for clonal neoantigens. T cells recognizing clonal neoantigens were detectable in patients with durable clinical benefit. Cytotoxic chemotherapy–induced subclonal neoantigens, contributing to an increased mutational load, were enriched in certain poor responders. These data suggest that neoantigen heterogeneity may influence immune surveillance and support therapeutic developments targeting clonal neoantigens. PMID:26940869

  3. Mitosis-specific phosphorylation of PML at T409 regulates spindle checkpoint.

    Science.gov (United States)

    Jin, J; Liu, J

    2016-08-31

    During mitosis, Promyelocytic leukemia nuclear bodies (PML NBs) change dramatically in morphology and composition, but little is known about function of PML in mitosis. Here, we show that PML is phosphorylated at T409 (PML p409) in a mitosis-specific manner. More importantly, PML p409 contributes to maintain the duration of pro-metaphase and regulates spindle checkpoint. Deficient PML p409 caused a shortening of pro-metaphase and challenged the nocodazole-triggered mitotic arrest. T409A mutation led to a higher frequency of misaligned chromosomes on metaphase plate, and subsequently death in late mitosis. In addition, inhibition of PML p409 repressed growth of tumor cells, suggesting that PML p409 is a potential target for cancer therapy. Collectively, our study demonstrated an important phosphorylated site of PML, which contributed to explore the role of PML in mitosis.

  4. A direct role of Mad1 in the spindle assembly checkpoint beyond Mad2 kinetochore recruitment

    DEFF Research Database (Denmark)

    Kruse, Thomas; Larsen, Marie Sofie Yoo; Sedgwick, Garry G

    2014-01-01

    in the SAC beyond recruitment of C-Mad2 to kinetochores has not yet been addressed. Here, we show that Mad1 is required for mitotic arrest even when C-Mad2 is artificially recruited to kinetochores, indicating that it has indeed an additional function in promoting the checkpoint. The C-terminal globular...... domain of Mad1 and conserved residues in this region are required for this unexpected function of Mad1........ The conversion of O-Mad2 into C-Mad2 at unattached kinetochores is thought to be a key step in activating the SAC. The "template model" proposes that this is achieved by the recruitment of soluble O-Mad2 to C-Mad2 bound at kinetochores through its interaction with Mad1. Whether Mad1 has additional roles...

  5. The Aurora B kinase in chromosome biorientation and spindle checkpoint signalling

    Directory of Open Access Journals (Sweden)

    Veronica eKrenn

    2015-10-01

    Full Text Available Aurora B, a member of the Aurora family of serine/threonine protein kinases, is a key player in chromosome segregation. As part of a macromolecular complex known as the chromosome passenger complex, Aurora B concentrates early during mitosis in the proximity of centromeres and kinetochores, the sites of attachment of chromosomes to spindle microtubules. There, it contributes to a number of processes that impart fidelity to cell division, including kinetochore stabilization, kinetochore-microtubule attachment, and the regulation of a surveillance mechanism named the spindle assembly checkpoint. In the regulation of these processes, Aurora B is the fulcrum of a remarkably complex network of interactions that feed back on its localization and activation state. In this review we discuss the multiple roles of Aurora B during mitosis, focusing in particular on its role at centromeres and kinetochores. Many details of the network of interactions at these locations remain poorly understood, and we focus here on several crucial outstanding questions.

  6. Arabidopsis ARGONAUTE7 selects miR390 through multiple checkpoints during RISC assembly.

    Science.gov (United States)

    Endo, Yayoi; Iwakawa, Hiro-oki; Tomari, Yukihide

    2013-07-01

    Plant ARGONAUTE7 (AGO7) assembles RNA-induced silencing complex (RISC) specifically with miR390 and regulates the auxin-signalling pathway via production of TAS3 trans-acting siRNAs (tasiRNAs). However, how AGO7 discerns miR390 among other miRNAs remains unclear. Here, we show that the 5' adenosine of miR390 and the central region of miR390/miR390* duplex are critical for the specific interaction with AGO7. Furthermore, despite the existence of mismatches in the seed and central regions of the duplex, cleavage of the miR390* strand is required for maturation of AGO7-RISC. These findings suggest that AGO7 uses multiple checkpoints to select miR390, thereby circumventing promiscuous tasiRNA production.

  7. Immune checkpoint inhibitors and targeted therapies for metastatic melanoma: A network meta-analysis.

    Science.gov (United States)

    Pasquali, Sandro; Chiarion-Sileni, Vanna; Rossi, Carlo Riccardo; Mocellin, Simone

    2017-03-01

    Immune checkpoint inhibitors and targeted therapies, two new class of drugs for treatment of metastatic melanoma, have not been compared in randomized controlled trials (RCT). We quantitatively summarized the evidence and compared immune and targeted therapies in terms of both efficacy and toxicity. A comprehensive search for RCTs of immune checkpoint inhibitors and targeted therapies was conducted to August 2016. Using a network meta-analysis approach, treatments were compared with each other and ranked based on their effectiveness (as measured by the impact on progression-free survival [PFS]) and acceptability (the inverse of high grade toxicity). Twelve RCTs enrolling 6207 patients were included. Network meta-analysis generated 15 comparisons. Combined BRAF and MEK inhibitors were associated with longer PFS as compared to anti-CTLA4 (HR: 0.22; 95% confidence interval [CI]: 0.12-0.41) and anti-PD1 antibodies alone (HR: 0.38; CI: 0.20-0.72). However, anti-PD1 monoclonal antibodies were less toxic than anti-CTLA4 monoclonal antibodies (RR: 0.65; CI: 0.40-0.78) and their combination significantly increased toxicity compared to either single agent anti-CTLA4 (RR: 2.06; CI: 1.45-2.93) or anti-PD1 monoclonal antibodies (RR: 3.67; CI: 2.27-5.96). Consistently, ranking analysis suggested that the combination of targeted therapies is the most effective strategy, whereas single agent anti-PD1 antibodies have the best acceptability. The GRADE level of evidence quality for these findings was moderate to low. The simultaneous inhibition of BRAF and MEK appears the most effective treatment for melanomas harboring BRAF V600 mutation, although anti-PD1 antibodies appear to be less toxic. Further research is needed to increase the quality of evidence. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. An Archaeosome-Adjuvanted Vaccine and Checkpoint Inhibitor Therapy Combination Significantly Enhances Protection from Murine Melanoma

    Directory of Open Access Journals (Sweden)

    Felicity C. Stark

    2017-10-01

    Full Text Available Archaeosomes constitute archaeal lipid vesicle vaccine adjuvants that evoke a strong CD8+ T cell response to antigenic cargo. Therapeutic treatment of murine B16-ovalbumin (B16-OVA melanoma with archaeosome-OVA eliminates small subcutaneous solid tumors; however, they eventually resurge despite an increased frequency of circulating and tumor infiltrating OVA-CD8+ T cells. Herein, a number of different approaches were evaluated to improve responses, including dose number, interval, and the combination of vaccine with checkpoint inhibitors. Firstly, we found that tumor protection could not be enhanced by repetitive and/or delayed boosting to maximize the CD8+ T cell number and/or phenotype. The in vivo cytotoxicity of vaccine-induced OVA-CD8+ T cells was impaired in tumor-bearing mice. Additionally, tumor-infiltrating OVA-CD8+ T cells had an increased expression of programmed cell death protein-1 (PD-1 compared to other organ compartments, suggesting impaired function. Combination therapy of tumor-bearing mice with the vaccine archaeosome-OVA, and α-CTLA-4 administered concurrently as well as α-PD-1 and an α-PD-L1 antibody administered starting 9 days after tumor challenge given on a Q3Dx4 schedule (days 9, 12, 15 and 18, significantly enhanced survival. Following multi-combination therapy ~70% of mice had rapid tumor recession, with no detectable tumor mass after >80 days in comparison to a median survival of 17–22 days for untreated or experimental groups receiving single therapies. Overall, archaeosomes offer a powerful platform for delivering cancer antigens when used in combination with checkpoint inhibitor immunotherapies.

  9. Damaged Skylab

    Science.gov (United States)

    1973-01-01

    The Saturn V vehicle, carrying the unmarned orbital workshop for the Skylab-1 mission, lifted off successfully and all systems performed normally. Sixty-three seconds into the flight, engineers in the operation support and control center saw an unexpected telemetry indication that signalled that damages occurred on one solar array and the micrometeoroid shield during the launch. The micrometeoroid shield, a thin protective cylinder surrounding the workshop protecting it from tiny space particles and the sun's scorching heat, ripped loose from its position around the workshop. This caused the loss of one solar wing and jammed the other. Still unoccupied, the Skylab was stricken with the loss of the heat shield and sunlight beat mercilessly on the lab's sensitive skin. Internal temperatures soared, rendering the station uninhabitable, threatening foods, medicines, films, and experiments. This image, taken during a fly-around inspection by the Skylab-2 crew, shows a crippled Skylab in orbit. The crew found their home in space to be in serious shape; the heat shield gone, one solar wing gone, and the other jammed. The Marshall Space Flight Center (MSFC) developed, tested, rehearsed, and approved three repair options. These options included a parasol sunshade and a twin-pole sunshade to restore the temperature inside the workshop, and a set of metal cutting tools to free the jammed solar panel.

  10. Structural damage

    International Nuclear Information System (INIS)

    Gray, R.E.; Bruhn, R.W.

    1992-01-01

    Virtually all structures show some signs of distress due to deterioration of the building components, to changed loads, or to changed support conditions. Changed support conditions result from ground movements. In mining regions many cases of structural distress are attributed to mining without considering alternative causes. This is particularly true of coal mining since it occurs under extensive areas. Coal mining is estimated to have already undermined more than eight million acres and may eventually undermine 40 million acres in the United States. Other nonmetal and metal underground mines impact much smaller areas. Although it is sometimes difficult, even with careful study, to identify the actual cause of damage, persons responsible for underground coal mining should at least be aware of possible causes of building stress other than mine subsidence. This paper presents information on distress to structures and briefly reviews a number of causes of ground movements other than subsidence: Mass movements, dissolution, erosion, frost action, shrinking and swelling, yield into excavations and compressibility

  11. Radiation damage prediction system using damage function

    International Nuclear Information System (INIS)

    Tanaka, Yoshihisa; Mori, Seiji

    1979-01-01

    The irradiation damage analysis system using a damage function was investigated. This irradiation damage analysis system consists of the following three processes, the unfolding of a damage function, the calculation of the neutron flux spectrum of the object of damage analysis and the estimation of irradiation effect of the object of damage analysis. The damage function is calculated by applying the SAND-2 code. The ANISN and DOT3, 5 codes are used to calculate neutron flux. The neutron radiation and the allowable time of reactor operation can be estimated based on these calculations of the damage function and neutron flux. The flow diagram of the process of analyzing irradiation damage by a damage function and the flow diagram of SAND-2 code are presented, and the analytical code for estimating damage, which is determined with a damage function and a neutron spectrum, is explained. The application of the irradiation damage analysis system using a damage function was carried out to the core support structure of a fast breeder reactor for the damage estimation and the uncertainty evaluation. The fundamental analytical conditions and the analytical model for this work are presented, then the irradiation data for SUS304, the initial estimated values of a damage function, the error analysis for a damage function and the analytical results are explained concerning the computation of a damage function for 10% total elongation. Concerning the damage estimation of FBR core support structure, the standard and lower limiting values of damage, the permissible neutron flux and the allowable years of reactor operation are presented and were evaluated. (Nakai, Y.)

  12. The healing of alkali-injured cornea is stimulated by a novel matrix regenerating agent (RGTA, CACICOL20): a biopolymer mimicking heparan sulfates reducing proteolytic, oxidative and nitrosative damage.

    Science.gov (United States)

    Cejkova, Jitka; Olmiere, Celine; Cejka, Cestmir; Trosan, Peter; Holan, Vladimir

    2014-04-01

    The efficacy of a chemically modified dextran - heparan sulfate mimicking regenerating agent (RGTA) on the healing of the rabbit cornea injured with alkali was examined. The eyes were injured with 0.15 N NaOH applied on the cornea or with 1.0 N NaOH using a 8 mm diameter filter paper disk. Then RGTA or placebo was applied on the cornea. In the last group of rabbits, corneas injured with the high alkali concentration were left without any treatment for four weeks; subsequently, the corneas were treated with RGTA or placebo. The central corneal thickness was measured using a pachymeter. The corneas were examined morphologically, immunohistochemically and for real time-PCR. Compared to control (unaffected) corneas, following the application of low alkali concentration the expression of urokinase-type plasminogen activator, metalloproteinase 9, nitric oxide synthase and xanthine oxidase was increased in the injured corneal epithelium of placebo-treated eyes, whereas the expression of antioxidant enzymes was reduced. Nitrotyrosine and malondialdehyde stainings appeared in the corneal epithelium. RGTA application suppressed the antioxidant/prooxidant imbalance and reduced the expression of the above-mentioned immunohistochemical markers. The corneal thickness increased after alkali injury, decreased during corneal healing after RGTA treatment faster than after placebo application. Following the injury with the high alkali concentration, corneal inflammation and neovascularization were highly pronounced in placebo-treated corneas, whereas in RGTA-treated corneas they were significantly supressed. When RGTA or placebo application was started later after alkali injury and corneas were ulcerated, subsequent RGTA treatment healed the majority of them. In conclusion, RGTA facilitates the healing of injured corneas via a reduction of proteolytic, oxidative and nitrosative damage.

  13. Chemotherapeutic Drugs: DNA Damage and Repair in Glioblastoma.

    Science.gov (United States)

    Annovazzi, Laura; Mellai, Marta; Schiffer, Davide

    2017-05-26

    Despite improvements in therapeutic strategies, glioblastoma (GB) remains one of the most lethal cancers. The presence of the blood-brain barrier, the infiltrative nature of the tumor and several resistance mechanisms account for the failure of current treatments. Distinct DNA repair pathways can neutralize the cytotoxicity of chemo- and radio-therapeutic agents, driving resistance and tumor relapse. It seems that a subpopulation of stem-like cells, indicated as glioma stem cells (GSCs), is responsible for tumor initiation, maintenance and recurrence and they appear to be more resistant owing to their enhanced DNA repair capacity. Recently, attention has been focused on the pivotal role of the DNA damage response (DDR) in tumorigenesis and in the modulation of therapeutic treatment effects. In this review, we try to summarize the knowledge concerning the main molecular mechanisms involved in the removal of genotoxic lesions caused by alkylating agents, emphasizing the role of GSCs. Beside their increased DNA repair capacity in comparison with non-stem tumor cells, GSCs show a constitutive checkpoint expression that enables them to survive to treatments in a quiescent, non-proliferative state. The targeted inhibition of checkpoint/repair factors of DDR can contribute to eradicate the GSC population and can have a great potential therapeutic impact aiming at sensitizing malignant gliomas to treatments, improving the overall survival of patients.

  14. The subunits of the S-phase checkpoint complex Mrc1/Tof1/Csm3: dynamics and interdependence.

    Science.gov (United States)

    Uzunova, Sonya Dimitrova; Zarkov, Alexander Stefanov; Ivanova, Anna Marianova; Stoynov, Stoyno Stefanov; Nedelcheva-Veleva, Marina Nedelcheva

    2014-01-01

    The S-phase checkpoint aims to prevent cells from generation of extensive single-stranded DNA that predisposes to genome instability. The S. cerevisiae complex Tof1/Csm3/Mrc1 acts to restrain the replicative MCM helicase when DNA synthesis is prohibited. Keeping the replication machinery intact allows restart of the replication fork when the block is relieved. Although the subunits of the Tof1/Csm3/Mrc1 complex are well studied, the impact of every single subunit on the triple complex formation and function needs to be established. This work studies the cellular localization and the chromatin binding of GFP-tagged subunits when the complex is intact and when a subunit is missing. We demonstrate that the complex is formed in cell nucleus, not the cytoplasm, as Tof1, Csm3 and Mrc1 enter the nucleus independently from one another. Via in situ chromatin binding assay we show that a Tof1-Csm3 dimer formation and chromatin binding is required to ensure the attachment of Mrc1 to chromatin. Our study indicates that the translocation into the nucleus is not the process to regulate the timing of chromatin association of Mrc1. We also studied the nuclear behavior of Mrc1 subunit in the process of adaptation to the presence hydroxyurea. Our results indicate that after prolonged HU incubation, cells bypass the S-phase checkpoint and proceed throughout the cell cycle. This process is accompanied by Mrc1 chromatin detachment and Rad53 dephosphorylation. In S. cerevisiae the subunits of the S-phase checkpoint complex Mrc1/Tof1/Csm3 independently enter the cell nucleus, where a Tof1-Csm3 dimer is formed to ensure the chromatin binding of Mrc1 and favor DNA replication and S-phase checkpoint fork arrest. In the process of adaptation to the presence of hydroxyurea Mrc1 is detached from chromatin and Rad53 checkpoint activity is diminished in order to allow S-phase checkpoint escape and completion of the cell cycle.

  15. Comparative Analysis of Immune Checkpoint Molecules and Their Potential Role in the Transmissible Tasmanian Devil Facial Tumor Disease

    Directory of Open Access Journals (Sweden)

    Andrew S. Flies

    2017-05-01

    Full Text Available Immune checkpoint molecules function as a system of checks and balances that enhance or inhibit immune responses to infectious agents, foreign tissues, and cancerous cells. Immunotherapies that target immune checkpoint molecules, particularly the inhibitory molecules programmed cell death 1 and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4, have revolutionized human oncology in recent years, yet little is known about these key immune signaling molecules in species other than primates and rodents. The Tasmanian devil facial tumor disease is caused by transmissible cancers that have resulted in a massive decline in the wild Tasmanian devil population. We have recently demonstrated that the inhibitory checkpoint molecule PD-L1 is upregulated on Tasmanian devil (Sarcophilus harrisii facial tumor cells in response to the interferon-gamma cytokine. As this could play a role in immune evasion by tumor cells, we performed a thorough comparative analysis of checkpoint molecule protein sequences among Tasmanian devils and eight other species. We report that many of the key signaling motifs and ligand-binding sites in the checkpoint molecules are highly conserved across the estimated 162 million years of evolution since the last common ancestor of placental and non-placental mammals. Specifically, we discovered that the CTLA-4 (MYPPPY ligand-binding motif and the CTLA-4 (GVYVKM inhibitory domain are completely conserved across all nine species used in our comparative analysis, suggesting that the function of CTLA-4 is likely conserved in these species. We also found that cysteine residues for intra- and intermolecular disulfide bonds were also highly conserved. For instance, all 20 cysteine residues involved in disulfide bonds in the human 4-1BB molecule were also present in devil 4-1BB. Although many key sequences were conserved, we have also identified immunoreceptor tyrosine-based inhibitory motifs (ITIMs and immunoreceptor tyrosine-based switch

  16. Clinical features, predictive correlates, and pathophysiology of immune-related adverse events in immune checkpoint inhibitor treatments in cancer: a short review

    OpenAIRE

    Yoest, Jennifer M

    2017-01-01

    Jennifer M Yoest Department of Pathology, University of Pittsburgh Medical Center, Pittsburgh, PA, USA Abstract: Identification and characterization of T-cell regulatory mechanisms, or checkpoints, have led to a wave of drug development aimed at inhibiting these targets to “remove the brakes” of the immune system. This class of anticancer therapeutics, termed immune checkpoint inhibitors (ICIs), has harnessed the potential of the body’s own immune system to reco...

  17. Using ergonomics checkpoints to support a participatory ergonomics intervention in an industrially developing country (IDC)--a case study.

    Science.gov (United States)

    Helali, Faramarz

    2009-01-01

    To achieve ergonomics awareness in 3 subsidiary companies, an intervention team was formed. The aims of this study were to implement basic ergonomics through a participatory ergonomics intervention process that can support a continuous learning process and lead to an improvement in health and safety as well as in the work systems in the organization. The findings of this study (i.e., method, continuous learning and integration) were key to making the participatory ergonomics intervention successful. Furthermore, 4 issues of the ergonomics checkpoints (i.e., work schedules, work tasks, healthy work organization and learning) for assessing the work system were found suitable for both changing work schedules and for improving the work system. This paper describes the result of this project and also the experiences gained and the conclusions reached from using the International Labour Office's ergonomics checkpoints in the industries of industrially developing country.

  18. Pharmacometrics and systems pharmacology of immune checkpoint inhibitor nivolumab in cancer translational medicine

    Directory of Open Access Journals (Sweden)

    Sujit Nair

    2016-02-01

    Full Text Available Nivolumab, a fully human immunoglobulin G4 (IgG4 monoclonal antibody (mAb that targets the programmed cell death-1 (PD-1 inhibitory receptor expressed on lymphocytes and dendritic cells, has been approved for metastatic melanoma, advanced squamous non-small cell lung cancer (NSCLC and metastatic renal cell carcinoma. In this review, pharmacology and pharmacometrics systems of this immunopharmaceutical are discussed. Mechanistic actions of T-cell biology with respect to both “priming phase” (anti-cytotoxic T-lymphocyte associated antigen 4 (anti-CTLA-4 mAb; ipilimumab and “effector phase” (anti-PD-1 mAb; nivolumab was discussed, respectively. Key pharmacometric variables in anticancer efficacy of nivolumab such as target engagement, metabolism, pharmacology systems and clearance are elucidated with an emphasis on current knowledge from pre-clinical as well as phase 1, 2 and 3 clinical trials information, including the data presented at the American Society of Clinical Oncology (ASCO 2015 and European Cancer Congress 2015. Nivolumab biomarkers, safety, and synergistic combination immunotherapies are delineated. Nivolumab, administered via intravenous infusion, has an acceptable safety profile and good efficacy. Indeed, the way forward to leverage maximum benefits for the cancer patient may be to synergize anti-PD-1 blockade with complementary targets in immune checkpoint pathways or other oncogenic signal transduction pathways. The encouraging results with nivolumab lend credence to the promise of immune checkpoint blockade as a therapeutic strategy that has been come-of-age in clinical oncology. Of necessity, the burden of “financial toxicity” on cancer patients and families must be factored in considering nivolumab therapy. The problem of ligand PD-L1 being a weak biomarker in clinical practice was discussed. Appropriate patient selection methods including immunopharmacogenomics may be used to identify those patients who are most

  19. Downregulation of Wip1 phosphatase modulates the cellular threshold of DNA damage signaling in mitosis

    Science.gov (United States)

    Macurek, Libor; Benada, Jan; Müllers, Erik; Halim, Vincentius A.; Krejčíková, Kateřina; Burdová, Kamila; Pecháčková, Sona; Hodný, Zdeněk; Lindqvist, Arne; Medema, René H.; Bartek, Jiri

    2013-01-01

    Cells are constantly challenged by DNA damage and protect their genome integrity by activation of an evolutionary conserved DNA damage response pathway (DDR). A central core of DDR is composed of a spatiotemporally ordered net of post-translational modifications, among which protein phosphorylation plays a major role. Activation of checkpoint kinases ATM/ATR and Chk1/2 leads to a temporal arrest in cell cycle progression (checkpoint) and allows time for DNA repair. Following DNA repair, cells re-enter the cell cycle by checkpoint recovery. Wip1 phosphatase (also called PPM1D) dephosphorylates multiple proteins involved in DDR and is essential for timely termination of the DDR. Here we have investigated how Wip1 is regulated in the context of the cell cycle. We found that Wip1 activity is downregulated by several mechanisms during mitosis. Wip1 protein abundance increases from G1 phase to G2 and declines in mitosis. Decreased abundance of Wip1 during mitosis is caused by proteasomal degradation. In addition, Wip1 is phosphorylated at multiple residues during mitosis, and this leads to inhibition of its enzymatic activity. Importantly, ectopic expression of Wip1 reduced γH2AX staining in mitotic cells and decreased the number of 53BP1 nuclear bodies in G1 cells. We propose that the combined decrease and inhibition of Wip1 in mitosis decreases the threshold necessary for DDR activation and enables cells to react adequately even to modest levels of DNA damage encountered during unperturbed mitotic progression. PMID:23255129

  20. Well screening for matrix stimulation treatments

    International Nuclear Information System (INIS)

    Saavedra, N; Solano, R; Gidley, J; Reyes, C.A; Rodriguez; Kondo, F; Hernandez, J

    1998-01-01

    Matrix acidizing is a stimulation technique only applicable to wells with surrounding damage. It is therefore very important to differentiate the real formation damage from the damage caused by flow Ni dynamic effects. The mechanical damage corresponds to flow restrictions caused by partial penetration, poor perforation as well as to reduce diameters of the production tubing. The dynamic effects are generated by inertia caused by high flow rates and high-pressure differentials. A common practice in our oil fields is to use a general formulation as acid treatment, most of the times without previous lab studies that guarantee the applicability of the treatment in the formation. Additionally, stimulation is randomly applied even treating undamaged wells with negative results and in the best of the cases, loss of the treatment. The selection of the well for matrix stimulation is an essential factor for the success of the treatment. Selection is done through the evaluation of the skin factor (S) and of the economic benefits of reducing the skin in comparison to the cost of the work. The most appropriate tool for skin evaluation is a good pressure test where the radial flow period can be identified. Nevertheless, we normally find-outdated tests most of the times taken with inaccurate tools. The interpretation problem is worsened by completions in which there is simultaneous production from several sand packages and it is difficult to individually differentiate damage factors. This works states a procedure for the selection of wells appropriate for stimulation; it also proposes a method to evaluate the skin factor when there are no accurate interpretations of the pressure tests. A new and increasingly applied methodology to treat wells with high water cuts, which are usually discarded due to the risk of stimulating water zones, is also mentioned

  1. Immune Checkpoint Inhibitor-Associated Type 1 Diabetes Mellitus: Case Series, Review of the Literature, and Optimal Management

    OpenAIRE

    Kapke, Jonathan; Shaheen, Zachary; Kilari, Deepak; Knudson, Paul; Wong, Stuart

    2017-01-01

    With the introduction of immune checkpoint inhibitors into clinical practice, various autoimmune toxicities have been described. Antibodies targeting the receptor:ligand pairing of programmed death receptor-1 (PD-1) and its cognate ligand programmed death-ligand 1 (PD-L1) in rare reports have been associated with autoimmune diabetes mellitus. We report 2 cases of rapid-onset, insulin-dependent, type 1 diabetes mellitus in the setting of administration of nivolumab, a fully human monoclonal an...

  2. Clinical characteristics of patient selection and imaging predictors of outcome in solid tumors treated with checkpoint-inhibitors

    International Nuclear Information System (INIS)

    Rossi, Sabrina; Toschi, Luca; Castello, Angelo; Grizzi, Fabio; Mansi, Luigi; Lopci, Egesta

    2017-01-01

    The rapidly evolving knowledge on tumor immunology and the continuous implementation of immunotherapy in cancer have recently led to the FDA and EMA approval of several checkpoint inhibitors as immunotherapic agents in clinical practice. Anti-CTLA-4, anti-PD-1, and anti-PDL-1 antibodies are becoming standard of care in advanced melanoma, as well as in relapsed or metastatic lung and kidney cancer, demonstrating higher and longer response compared to standard chemotherapy. These encouraging results have fostered the evaluation of these antibodies either alone or in combination with other therapies in several dozen clinical trials for the treatment of multiple tumor types. However, not all patients respond to immune checkpoint inhibitors, hence, specific biomarkers are necessary to guide and monitor therapy. The utility of PD-L1 expression as a biomarker has varied in different clinical trials, but, to date, no consensus has been reached on whether PD-L1 expression is an ideal marker for response and patient selection; approximately 20-25% of patients will respond to immunotherapy with checkpoint inhibitors despite a negative PD-L1 staining. On the other hand, major issues concern the evaluation of objective response in patients treated with immunotherapy. Pure morphological criteria as commonly used in solid tumors (i.e. RECIST) are not sufficient because change in size is not an early and reliable marker of tumor response to biological therapies. Thus, the scientific community has required a continuous adaptation of immune-related response criteria (irRC) to overcome the problem. In this context, metabolic information and antibody-based imaging with positron emission tomography (PET) have been investigated, providing a powerful approach for an optimal stratification of patients at staging and during the evaluation of the response to therapy. In the present review we provide an overview on the clinical characteristics of patient selection when using imaging

  3. Clinical characteristics of patient selection and imaging predictors of outcome in solid tumors treated with checkpoint-inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Rossi, Sabrina; Toschi, Luca [Humanitas Clinical and Research Hospital, Medical Oncology, Rozzano (Italy); Castello, Angelo [Humanitas Clinical and Research Hospital, Nuclear Medicine, Rozzano (Italy); Grizzi, Fabio [Humanitas Clinical and Research Hospital, Immunology and Inflammation, Rozzano (Italy); Mansi, Luigi [Seconda Universita di Napoli, Nuclear Medicine, Naples (Italy); Lopci, Egesta [Humanitas Clinical and Research Hospital, Nuclear Medicine, Rozzano (Italy); Humanitas Cancer Center, Humanitas Clinical and Research Hospital, Nuclear Medicine, Rozzano, MI (Italy)

    2017-12-15

    The rapidly evolving knowledge on tumor immunology and the continuous implementation of immunotherapy in cancer have recently led to the FDA and EMA approval of several checkpoint inhibitors as immunotherapic agents in clinical practice. Anti-CTLA-4, anti-PD-1, and anti-PDL-1 antibodies are becoming standard of care in advanced melanoma, as well as in relapsed or metastatic lung and kidney cancer, demonstrating higher and longer response compared to standard chemotherapy. These encouraging results have fostered the evaluation of these antibodies either alone or in combination with other therapies in several dozen clinical trials for the treatment of multiple tumor types. However, not all patients respond to immune checkpoint inhibitors, hence, specific biomarkers are necessary to guide and monitor therapy. The utility of PD-L1 expression as a biomarker has varied in different clinical trials, but, to date, no consensus has been reached on whether PD-L1 expression is an ideal marker for response and patient selection; approximately 20-25% of patients will respond to immunotherapy with checkpoint inhibitors despite a negative PD-L1 staining. On the other hand, major issues concern the evaluation of objective response in patients treated with immunotherapy. Pure morphological criteria as commonly used in solid tumors (i.e. RECIST) are not sufficient because change in size is not an early and reliable marker of tumor response to biological therapies. Thus, the scientific community has required a continuous adaptation of immune-related response criteria (irRC) to overcome the problem. In this context, metabolic information and antibody-based imaging with positron emission tomography (PET) have been investigated, providing a powerful approach for an optimal stratification of patients at staging and during the evaluation of the response to therapy. In the present review we provide an overview on the clinical characteristics of patient selection when using imaging

  4. Computational analysis of transcranial magnetic stimulation in the presence of deep brain stimulation probes

    Science.gov (United States)

    Syeda, F.; Holloway, K.; El-Gendy, A. A.; Hadimani, R. L.

    2017-05-01

    Transcranial Magnetic Stimulation is an emerging non-invasive treatment for depression, Parkinson's disease, and a variety of other neurological disorders. Many Parkinson's patients receive the treatment known as Deep Brain Stimulation, but often require additional therapy for speech and swallowing impairment. Transcranial Magnetic Stimulation has been explored as a possible treatment by stimulating the mouth motor area of the brain. We have calculated induced electric field, magnetic field, and temperature distributions in the brain using finite element analysis and anatomically realistic heterogeneous head models fitted with Deep Brain Stimulation leads. A Figure of 8 coil, current of 5000 A, and frequency of 2.5 kHz are used as simulation parameters. Results suggest that Deep Brain Stimulation leads cause surrounding tissues to experience slightly increased E-field (Δ Emax =30 V/m), but not exceeding the nominal values induced in brain tissue by Transcranial Magnetic Stimulation without leads (215 V/m). The maximum temperature in the brain tissues surrounding leads did not change significantly from the normal human body temperature of 37 °C. Therefore, we ascertain that Transcranial Magnetic Stimulation in the mouth motor area may stimulate brain tissue surrounding Deep Brain Stimulation leads, but will not cause tissue damage.

  5. The internal Cdc20 binding site in BubR1 facilitates both spindle assembly checkpoint signalling and silencing

    DEFF Research Database (Denmark)

    Lischetti, Tiziana; Zhang, Gang; Sedgwick, Garry G

    2014-01-01

    Improperly attached kinetochores activate the spindle assembly checkpoint (SAC) and by an unknown mechanism catalyse the binding of two checkpoint proteins, Mad2 and BubR1, to Cdc20 forming the mitotic checkpoint complex (MCC). Here, to address the functional role of Cdc20 kinetochore localization...... in the SAC, we delineate the molecular details of its interaction with kinetochores. We find that BubR1 recruits the bulk of Cdc20 to kinetochores through its internal Cdc20 binding domain (IC20BD). We show that preventing Cdc20 kinetochore localization by removal of the IC20BD has a limited effect...... on the SAC because the IC20BD is also required for efficient SAC silencing. Indeed, the IC20BD can disrupt the MCC providing a mechanism for its role in SAC silencing. We thus uncover an unexpected dual function of the second Cdc20 binding site in BubR1 in promoting both efficient SAC signalling and SAC...

  6. DNA damage assessment by visualization and quantification of DNA damage response

    International Nuclear Information System (INIS)

    Matsuda, Shun; Matsuda, Tomonari; Ikura, Tsuyoshi

    2017-01-01

    DNA damage response (DDR) carries out signal transduction for DNA repair, activation of cell cycle checkpoint, and apoptosis to maintain genome integrity, in response to DNA damage. Many proteins and their post-translational modifications participate in the process. Especially, S139-phosphorylated histone H2AX (γH2AX), which is formed by DNA double-strand breaks (DSBs), is an important factor to bring and retain other DDR proteins to DSB sites, Thus, γH2AX is used as a good indicator of DSBs in clinical study and pharmacology for efficacy evaluation of chemotherapy and radiotherapy, detection of precancerous regions, and others. In regulatory science, γH2AX is also a useful biomarker of genotoxicity of chemicals, since a wide range of genotoxic chemicals induce γH2AX. However, conventional detection methods of γH2AX absolutely require anti-γH2AX antibody whose staining is burdensome and time-consuming, and some of these methods are not so superior in quantitativity. In this review, we introduce two new methods to overcome these limitations, involving an easy-to-use genotoxicity assay using DDR-visualizing cells and an absolute quantification method of γH2AX using liquid chromatography-tandem mass spectrometry (LC/MS/MS). (author)

  7. Implications of Sensory Stimulation in Self-Destructive Behavior.

    Science.gov (United States)

    Edelson, Stephen M.

    1984-01-01

    The author extends the self stimulatory theory of self destructive behavior in autistic, schizophrenic, and mentally retarded individuals to suggest that damage of the skin's nerve structure lowers the tactile sensory threshold for physical input and enables individuals to obtain sensory stimulation by repeatedly depressing the damaged area. (CL)

  8. The telomeric protein TRF2 binds the ATM kinase and can inhibit the ATM-dependent DNA damage response.

    Directory of Open Access Journals (Sweden)

    Jan Karlseder

    2004-08-01

    Full Text Available The telomeric protein TRF2 is required to prevent mammalian telomeres from activating DNA damage checkpoints. Here we show that overexpression of TRF2 affects the response of the ATM kinase to DNA damage. Overexpression of TRF2 abrogated the cell cycle arrest after ionizing radiation and diminished several other readouts of the DNA damage response, including phosphorylation of Nbs1, induction of p53, and upregulation of p53 targets. TRF2 inhibited autophosphorylation of ATM on S1981, an early step in the activation of this kinase. A region of ATM containing S1981 was found to directly interact with TRF2 in vitro, and ATM immunoprecipitates contained TRF2. We propose that TRF2 has the ability to inhibit ATM activation at telomeres. Because TRF2 is abundant at chromosome ends but not elsewhere in the nucleus, this mechanism of checkpoint control could specifically block a DNA damage response at telomeres without affecting the surveillance of chromosome internal damage.

  9. A comprehensive complex systems approach to the study and analysis of mammalian cell cycle control system in the presence of DNA damage stress.

    Science.gov (United States)

    Abroudi, Ali; Samarasinghe, Sandhya; Kulasiri, Don

    2017-09-21

    Not many models of mammalian cell cycle system exist due to its complexity. Some models are too complex and hard to understand, while some others are too simple and not comprehensive enough. Moreover, some essential aspects, such as the response of G1-S and G2-M checkpoints to DNA damage as well as the growth factor signalling, have not been investigated from a systems point of view in current mammalian cell cycle models. To address these issues, we bring a holistic perspective to cell cycle by mathematically modelling it as a complex system consisting of important sub-systems that interact with each other. This retains the functionality of the system and provides a clearer interpretation to the processes within it while reducing the complexity in comprehending these processes. To achieve this, we first update a published ODE mathematical model of cell cycle with current knowledge. Then the part of the mathematical model relevant to each sub-system is shown separately in conjunction with a diagram of the sub-system as part of this representation. The model sub-systems are Growth Factor, DNA damage, G1-S, and G2-M checkpoint signalling. To further simplify the model and better explore the function of sub-systems, they are further divided into modules. Here we also add important new modules of: chk-related rapid cell cycle arrest, p53 modules expanded to seamlessly integrate with the rapid arrest module, Tyrosine phosphatase modules that activate Cyc_Cdk complexes and play a crucial role in rapid and delay arrest at both G1-S and G2-M, Tyrosine Kinase module that is important for inactivating nuclear transport of CycB_cdk1 through Wee1 to resist M phase entry, Plk1-Related module that is crucial in activating Tyrosine phosphatases and inactivating Tyrosine kinase, and APC-Related module to show steps in CycB degradation. This multi-level systems approach incorporating all known aspects of cell cycle allowed us to (i) study, through dynamic simulation of an ODE model

  10. Is there a role for immune checkpoint blockade with ipilimumab in prostate cancer?

    International Nuclear Information System (INIS)

    Cha, Edward; Small, Eric J

    2013-01-01

    Treatment for advanced prostate cancer has and will continue to grow increasingly complex, owing to the introduction of multiple new therapeutic approaches with the potential to substantially improve outcomes for this disease. Agents that modulate the patient's immune system to fight prostate cancer – immunotherapeutics – are among the most exciting of these new approaches. The addition of antigen-specific immunotherapy to the treatment of castration-resistant prostate cancer (CRPC) has paved the way for additional research that seeks to augment the activity of the immune system itself. The monoclonal antibody ipilimumab, approved in over 40 countries to treat advanced melanoma and currently under phase 2 and 3 investigation in prostate cancer, is thought to act by augmenting immune responses to tumors through blockade of cytotoxic T-lymphocyte antigen 4, an inhibitory immune checkpoint molecule. Ipilimumab has been studied in seven phase 1 and 2 clinical trials that evaluated various doses, schedules, and combinations across the spectrum of patients with advanced prostate cancer. The CRPC studies of ipilimumab to date suggest that the agent is active in prostate cancer as monotherapy or in combination with radiotherapy, docetaxel, or other immunotherapeutics, and that the adverse event profile is as expected given the safety data in advanced melanoma. The ongoing phase 3 program will further characterize the risk/benefit profile of ipilimumab in chemotherapy-naïve and -pretreated CRPC

  11. The Spindle Assembly Checkpoint in Arabidopsis Is Rapidly Shut Off during Severe Stress.

    Science.gov (United States)

    Komaki, Shinichiro; Schnittger, Arp

    2017-10-23

    The spindle assembly checkpoint (SAC) in animals and yeast assures equal segregation of chromosomes during cell division. The prevalent occurrence of polyploidy in flowering plants together with the observation that many plants can be readily forced to double their genomes by application of microtubule drugs raises the question of whether plants have a proper SAC. Here, we provide a functional framework of the core SAC proteins in Arabidopsis. We reveal that Arabidopsis will delay mitosis in a SAC-dependent manner if the spindle is perturbed. However, we also show that the molecular architecture of the SAC is unique in plants. Moreover, the SAC is short-lived and cannot stay active for more than 2 hr, after which the cell cycle is reset. This resetting opens the possibility for genome duplications and raises the hypothesis that a rapid termination of a SAC-induced mitotic arrest provides an adaptive advantage for plants impacting plant genome evolution. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Morphogenesis checkpoint kinase Swe1 is the executor of lipolysis-dependent cell-cycle progression.

    Science.gov (United States)

    Chauhan, Neha; Visram, Myriam; Cristobal-Sarramian, Alvaro; Sarkleti, Florian; Kohlwein, Sepp D

    2015-03-10

    Cell growth and division requires the precise duplication of cellular DNA content but also of membranes and organelles. Knowledge about the cell-cycle-dependent regulation of membrane and storage lipid homeostasis is only rudimentary. Previous work from our laboratory has shown that the breakdown of triacylglycerols (TGs) is regulated in a cell-cycle-dependent manner, by activation of the Tgl4 lipase by the major cyclin-dependent kinase Cdc28. The lipases Tgl3 and Tgl4 are required for efficient cell-cycle progression during the G1/S (Gap1/replication phase) transition, at the onset of bud formation, and their absence leads to a cell-cycle delay. We now show that defective lipolysis activates the Swe1 morphogenesis checkpoint kinase that halts cell-cycle progression by phosphorylation of Cdc28 at tyrosine residue 19. Saturated long-chain fatty acids and phytosphingosine supplementation rescue the cell-cycle delay in the Tgl3/Tgl4 lipase-deficient strain, suggesting that Swe1 activity responds to imbalanced sphingolipid metabolism, in the absence of TG degradation. We propose a model by which TG-derived sphingolipids are required to activate the protein phosphatase 2A (PP2A(Cdc55)) to attenuate Swe1 phosphorylation and its inhibitory effect on Cdc28 at the G1/S transition of the cell cycle.

  13. Evaluation of response to immune checkpoint inhibitors: Is there a role for positron emission tomography?

    Institute of Scientific and Technical Information of China (English)

    Matteo Bauckneht; Roberta Piva; Gianmario Sambuceti; Francesco Grossi; Silvia Morbelli

    2017-01-01

    Strategies targeting intracellular negative regulators such as immune checkpoint inhibitors(ICPIs) have demonstrated significant antitumor activity across a wide range of solid tumors. In the clinical practice, the radiological effect of immunotherapeutic agents has raised several more relevant and complex challenges for the determination of their imaging-based response at single patient level. Accordingly, it has been suggested that the conventional Response Evaluation Criteria in Solid Tumors assessment alone, based on dimensional evaluation provided by computed tomography(CT), tends to underestimate the benefit of ICPIs at least in a subset of patients, supporting the need of immunerelated response criteria. Different from CT, very few data are available for the evaluation of immunotherapy by means of 18F-fluoro-2-deoxy-D-glucose positron emission tomography(FDG-PET). Moreover, since the antineoplastic activity of ICPIs is highly related to the activation of T cells against cancer cells, FDG accumulation might cause false-positive findings. Yet, discrimination between benign and malignant processes represents a huge challenge for FDG-PET in this clinical setting. Consequently, it might be of high interest to test the complex and variegated response to ICPIs by means of PET and thus it is worthwhile to ask if a similar introduction of immune-related PET-based criteria could be proposed in the future. Finally, PET might offer a new insight into the biology and pathophysiology of ICPIs thanks to a growing number of non-invasive immunediagnostic approaches based on non-FDG tracers.

  14. Inhibition of CDK7 bypasses spindle assembly checkpoint via premature cyclin B degradation during oocyte meiosis.

    Science.gov (United States)

    Wang, HaiYang; Jo, Yu-Jin; Sun, Tian-Yi; Namgoong, Suk; Cui, Xiang-Shun; Oh, Jeong Su; Kim, Nam-Hyung

    2016-12-01

    To ensure accurate chromosome segregation, the spindle assembly checkpoint (SAC) delays anaphase onset by preventing the premature activation of anaphase-promoting complex/cyclosome (APC/C) until all kinetochores are attached to the spindle. Although an escape from mitosis in the presence of unsatisfied SAC has been shown in several cancer cells, it has not been reported in oocyte meiosis. Here, we show that CDK7 activity is required to prevent a bypass of SAC during meiosis I in mouse oocytes. Inhibition of CDK7 using THZ1 accelerated the first meiosis, leading to chromosome misalignment, lag of chromosomes during chromosome segregation, and a high incidence of aneuploidy. Notably, this acceleration occurred in the presence of SAC proteins including Mad2 and Bub3 at the kinetochores. However, inhibition of APC/C-mediated cyclin B degradation blocked the THZ1-induced premature polar body extrusion. Moreover, chromosomal defects mediated by THZ1 were rescued when anaphase onset was delayed. Collectively, our results show that CDK7 activity is required to prevent premature anaphase onset by suppressing the bypass of SAC, thus ensuring chromosome alignment and proper segregation. These findings reveal new roles of CDK7 in the regulation of meiosis in mammalian oocytes. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. Arabidopsis pdr2 reveals a phosphate-sensitive checkpoint in root development.

    Science.gov (United States)

    Ticconi, Carla A; Delatorre, Carla A; Lahner, Brett; Salt, David E; Abel, Steffen

    2004-03-01

    Plants have evolved complex strategies to maintain phosphate (Pi) homeostasis and to maximize Pi acquisition when the macronutrient is limiting. Adjustment of root system architecture via changes in meristem initiation and activity is integral to the acclimation process. However, the mechanisms that monitor external Pi status and interpret the nutritional signal remain to be elucidated. Here, we present evidence that the Pi deficiency response, pdr2, mutation disrupts local Pi sensing. The sensitivity and amplitude of metabolic Pi-starvation responses, such as Pi-responsive gene expression or accumulation of anthocyanins and starch, are enhanced in pdr2 seedlings. However, the most conspicuous alteration of pdr2 is a conditional short-root phenotype that is specific for Pi deficiency and caused by selective inhibition of root cell division followed by cell death below a threshold concentration of about 0.1 mm external Pi. Measurements of general Pi uptake and of total phosphorus (P) in root tips exclude a defect in high-affinity Pi acquisition. Rescue of root meristem activity in Pi-starved pdr2 by phosphite (Phi), a non-metabolizable Pi analog, and divided-root experiments suggest that pdr2 disrupts sensing of low external Pi availability. Thus, PDR2 is proposed to function at a Pi-sensitive checkpoint in root development, which monitors environmental Pi status, maintains and fine-tunes meristematic activity, and finally adjusts root system architecture to maximize Pi acquisition.

  16. Centromeric DNA replication reconstitution reveals DNA loops and ATR checkpoint suppression.

    Science.gov (United States)

    Aze, Antoine; Sannino, Vincenzo; Soffientini, Paolo; Bachi, Angela; Costanzo, Vincenzo

    2016-06-01

    Half of the human genome is made up of repetitive DNA. However, mechanisms underlying replication of chromosome regions containing repetitive DNA are poorly understood. We reconstituted replication of defined human chromosome segments using bacterial artificial chromosomes in Xenopus laevis egg extract. Using this approach we characterized the chromatin assembly and replication dynamics of centromeric alpha-satellite DNA. Proteomic analysis of centromeric chromatin revealed replication-dependent enrichment of a network of DNA repair factors including the MSH2-6 complex, which was required for efficient centromeric DNA replication. However, contrary to expectations, the ATR-dependent checkpoint monitoring DNA replication fork arrest could not be activated on highly repetitive DNA due to the inability of the single-stranded DNA binding protein RPA to accumulate on chromatin. Electron microscopy of centromeric DNA and supercoil mapping revealed the presence of topoisomerase I-dependent DNA loops embedded in a protein matrix enriched for SMC2-4 proteins. This arrangement suppressed ATR signalling by preventing RPA hyper-loading, facilitating replication of centromeric DNA. These findings have important implications for our understanding of repetitive DNA metabolism and centromere organization under normal and stressful conditions.

  17. The flavonoid eupatorin inactivates the mitotic checkpoint leading to polyploidy and apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Salmela, Anna-Leena [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Graduate School of Biomedical Sciences, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Pouwels, Jeroen; Kukkonen-Macchi, Anu [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Waris, Sinikka; Toivonen, Pauliina [Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Jaakkola, Kimmo [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Maeki-Jouppila, Jenni [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Drug Discovery Graduate School, University of Turku (Finland); Kallio, Lila, E-mail: lila.kallio@vtt.fi [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Kallio, Marko J. [VTT Technical Research Centre of Finland, Medical Biotechnology, P.O. Box 106, Turku (Finland); Turku Centre for Biotechnology, P.O. Box 123, University of Turku (Finland); Centre of Excellence for Translational Genome-Scale Biology, P.O. Box 106, Academy of Finland (Finland)

    2012-03-10

    The spindle assembly checkpoint (SAC) is a conserved mechanism that ensures the fidelity of chromosome distribution in mitosis by preventing anaphase onset until the correct bipolar microtubule-kinetochore attachments are formed. Errors in SAC function may contribute to tumorigenesis by inducing numerical chromosome anomalies (aneuploidy). On the other hand, total disruption of SAC can lead to massive genomic imbalance followed by cell death, a phenomena that has therapeutic potency. We performed a cell-based high-throughput screen with a compound library of 2000 bioactives for novel SAC inhibitors and discovered a plant-derived phenolic compound eupatorin (3 Prime ,5-dihydroxy-4 Prime ,6,7-trimethoxyflavone) as an anti-mitotic flavonoid. The premature override of the microtubule drug-imposed mitotic arrest by eupatorin is dependent on microtubule-kinetochore attachments but not interkinetochore tension. Aurora B kinase activity, which is essential for maintenance of normal SAC signaling, is diminished by eupatorin in cells and in vitro providing a mechanistic explanation for the observed forced mitotic exit. Eupatorin likely has additional targets since eupatorin treatment of pre-mitotic cells causes spindle anomalies triggering a transient M phase delay followed by impaired cytokinesis and polyploidy. Finally, eupatorin potently induces apoptosis in multiple cancer cell lines and suppresses cancer cell proliferation in organotypic 3D cell culture model.

  18. Dietary flavonoid fisetin induces a forced exit from mitosis by targeting the mitotic spindle checkpoint

    Science.gov (United States)

    Salmela, Anna-Leena; Pouwels, Jeroen; Varis, Asta; Kukkonen, Anu M.; Toivonen, Pauliina; Halonen, Pasi K.; Perälä, Merja; Kallioniemi, Olli; Gorbsky, Gary J.; Kallio, Marko J.

    2009-01-01

    Fisetin is a natural flavonol present in edible vegetables, fruits and wine at 2–160 μg/g concentrations and an ingredient in nutritional supplements with much higher concentrations. The compound has been reported to exert anticarcinogenic effects as well as antioxidant and anti-inflammatory activity via its ability to act as an inhibitor of cell proliferation and free radical scavenger, respectively. Our cell-based high-throughput screen for small molecules that override chemically induced mitotic arrest identified fisetin as an antimitotic compound. Fisetin rapidly compromised microtubule drug-induced mitotic block in a proteasome-dependent manner in several human cell lines. Moreover, in unperturbed human cancer cells fisetin caused premature initiation of chromosome segregation and exit from mitosis without normal cytokinesis. To understand the molecular mechanism behind these mitotic errors, we analyzed the consequences of fisetin treatment on the localization and phoshorylation of several mitotic proteins. Aurora B, Bub1, BubR1 and Cenp-F rapidly lost their kinetochore/centromere localization and others became dephosphorylated upon addition of fisetin to the culture medium. Finally, we identified Aurora B kinase as a novel direct target of fisetin. The activity of Aurora B was significantly reduced by fisetin in vitro and in cells, an effect that can explain the observed forced mitotic exit, failure of cytokinesis and decreased cell viability. In conclusion, our data propose that fisetin perturbs spindle checkpoint signaling, which may contribute to the antiproliferative effects of the compound. PMID:19395653

  19. Dual personality of Mad1: regulation of nuclear import by a spindle assembly checkpoint protein.

    Science.gov (United States)

    Cairo, Lucas V; Ptak, Christopher; Wozniak, Richard W

    2013-01-01

    Nuclear transport is a dynamic process that can be modulated in response to changes in cellular physiology. We recently reported that the transport activity of yeast nuclear pore complexes (NPCs) is altered in response to kinetochore-microtubule (KT-MT) interaction defects. Specifically, KT detachment from MTs activates a signaling pathway that prevents the nuclear import of cargos by the nuclear transport factor Kap121p. This loss of Kap121p-mediated import is thought to influence the nuclear environment, including the phosphorylation state of nuclear proteins. A key regulator of this process is the spindle assembly checkpoint protein Mad1p. In response to unattached KTs, Mad1p dynamically cycles between NPCs and KTs. This cycling appears to induce NPC molecular rearrangements that prevent the nuclear import of Kap121p-cargo complexes. Here, we discuss the underlying mechanisms and the physiological relevance of Mad1p cycling and the inhibition of Kap121p-mediated nuclear import, focusing on outstanding questions within the pathway.

  20. The bacterial cell cycle checkpoint protein Obg and its role in programmed cell death

    Directory of Open Access Journals (Sweden)

    Liselot Dewachter

    2016-03-01

    Full Text Available The phenomenon of programmed cell death (PCD, in which cells initiate their own demise, is not restricted to multicellular organisms. Unicellular organisms, both eukaryotes and prokaryotes, also possess pathways that mediate PCD. We recently identified a PCD mechanism in Escherichia coli that is triggered by a mutant isoform of the essential GTPase ObgE (Obg of E. coli. Importantly, the PCD pathway mediated by mutant Obg (Obg* differs fundamentally from other previously described bacterial PCD pathways and thus constitutes a new mode of PCD. ObgE was previously proposed to act as a cell cycle checkpoint protein able to halt cell division. The implication of ObgE in the regulation of PCD further increases the similarity between this protein and eukaryotic cell cycle regulators that are capable of doing both. Moreover, since Obg is conserved in eukaryotes, the elucidation of this cell death mechanism might contribute to the understanding of PCD in higher organisms. Additionally, if Obg*-mediated PCD is conserved among different bacterial species, it will be a prime target for the development of innovative antibacterials that artificially induce this pathway.

  1. Optimising measles virus-guided radiovirotherapy with external beam radiotherapy and specific checkpoint kinase 1 inhibition

    International Nuclear Information System (INIS)

    Touchefeu, Yann; Khan, Aadil A.; Borst, Gerben; Zaidi, Shane H.; McLaughlin, Martin; Roulstone, Victoria; Mansfield, David; Kyula, Joan; Pencavel, Tim; Karapanagiotou, Eleni M.; Clayton, Jamie; Federspiel, Mark J.; Russell, Steve J.; Garrett, Michelle; Collins, Ian; Harrington, Kevin J.

    2013-01-01

    Background and purpose: We previously reported a therapeutic strategy comprising replication-defective NIS-expressing adenovirus combined with radioiodide, external beam radiotherapy (EBRT) and DNA repair inhibition. We have now evaluated NIS-expressing oncolytic measles virus (MV-NIS) combined with NIS-guided radioiodide, EBRT and specific checkpoint kinase 1 (Chk1) inhibition in head and neck and colorectal models. Materials and methods: Anti-proliferative/cytotoxic effects of individual agents and their combinations were measured by MTS, clonogenic and Western analysis. Viral gene expression was measured by radioisotope uptake and replication by one-step growth curves. Potential synergistic interactions were tested in vitro by Bliss independence analysis and in in vivo therapeutic studies. Results: EBRT and MV-NIS were synergistic in vitro. Furthermore, EBRT increased NIS expression in infected cells. SAR-020106 was synergistic with EBRT, but also with MV-NIS in HN5 cells. MV-NIS mediated 131 I-induced cytotoxicity in HN5 and HCT116 cells and, in the latter, this was enhanced by SAR-020106. In vivo studies confirmed that MV-NIS, EBRT and Chk1 inhibition were effective in HCT116 xenografts. The quadruplet regimen of MV-NIS, virally-directed 131 I, EBRT and SAR-020106 had significant anti-tumour activity in HCT116 xenografts. Conclusion: This study strongly supports translational and clinical research on MV-NIS combined with radiation therapy and radiosensitising agents

  2. Drosophila Polo regulates the spindle assembly checkpoint through Mps1-dependent BubR1 phosphorylation.

    Science.gov (United States)

    Conde, Carlos; Osswald, Mariana; Barbosa, João; Moutinho-Santos, Tatiana; Pinheiro, Diana; Guimarães, Sofia; Matos, Irina; Maiato, Helder; Sunkel, Claudio E

    2013-06-12

    Maintenance of genomic stability during eukaryotic cell division relies on the spindle assembly checkpoint (SAC) that prevents mitotic exit until all chromosomes are properly attached to the spindle. Polo is a mitotic kinase proposed to be involved in SAC function, but its role has remained elusive. We demonstrate that Polo and Aurora B functional interdependency comprises a positive feedback loop that promotes Mps1 kinetochore localization and activity. Expression of constitutively active Polo restores normal Mps1 kinetochore levels even after Aurora B inhibition, highlighting a role for Polo in Mps1 recruitment to unattached kinetochores downstream of Aurora B. We also show that Mps1 kinetochore localization is required for BubR1 hyperphosphorylation and formation of the 3F3/2 phosphoepitope. This is essential to allow recruitment of Cdc20 to unattached kinetochores and the assembly of anaphase-promoting complex/cyclosome-inhibitory complexes to levels that ensure long-term SAC activity. We propose a model in which Polo controls Mps1-dependent BubR1 phosphorylation to promote Cdc20 kinetochore recruitment and sustained SAC function.

  3. Early programming and late-acting checkpoints governing the development of CD4 T cell memory.

    Science.gov (United States)

    Dhume, Kunal; McKinstry, K Kai

    2018-04-27

    CD4 T cells contribute to protection against pathogens through numerous mechanisms. Incorporating the goal of memory CD4 T cell generation into vaccine strategies thus offers a powerful approach to improve their efficacy, especially in situations where humoral responses alone cannot confer long-term immunity. These threats include viruses such as influenza that mutate coat proteins to avoid neutralizing antibodies, but that are targeted by T cells that recognize more conserved protein epitopes shared by different strains. A major barrier in the design of such vaccines is that the mechanisms controlling the efficiency with which memory cells form remain incompletely understood. Here, we discuss recent insights into fate decisions controlling memory generation. We focus on the importance of three general cues: interleukin-2, antigen, and costimulatory interactions. It is increasingly clear that these signals have a powerful influence on the capacity of CD4 T cells to form memory during two distinct phases of the immune response. First, through 'programming' that occurs during initial priming, and second, through 'checkpoints' that operate later during the effector stage. These findings indicate that novel vaccine strategies must seek to optimize cognate interactions, during which interleukin-2-, antigen, and costimulation-dependent signals are tightly linked, well beyond initial antigen encounter to induce robust memory CD4 T cells. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  4. Checkpoint Kinase Rad53 Couples Leading- and Lagging-Strand DNA Synthesis under Replication Stress.

    Science.gov (United States)

    Gan, Haiyun; Yu, Chuanhe; Devbhandari, Sujan; Sharma, Sushma; Han, Junhong; Chabes, Andrei; Remus, Dirk; Zhang, Zhiguo

    2017-10-19

    The checkpoint kinase Rad53 is activated during replication stress to prevent fork collapse, an essential but poorly understood process. Here we show that Rad53 couples leading- and lagging-strand synthesis under replication stress. In rad53-1 cells stressed by dNTP depletion, the replicative DNA helicase, MCM, and the leading-strand DNA polymerase, Pol ε, move beyond the site of DNA synthesis, likely unwinding template DNA. Remarkably, DNA synthesis progresses further along the lagging strand than the leading strand, resulting in the exposure of long stretches of single-stranded leading-strand template. The asymmetric DNA synthesis in rad53-1 cells is suppressed by elevated levels of dNTPs in vivo, and the activity of Pol ε is compromised more than lagging-strand polymerase Pol δ at low dNTP concentrations in vitro. Therefore, we propose that Rad53 prevents the generation of excessive ssDNA under replication stress by coordinating DNA unwinding with synthesis of both strands. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. G2 checkpoint abrogator abates the antagonistic interaction between antimicrotubule drugs and radiation therapy

    International Nuclear Information System (INIS)

    Sui Meihua; Zhang Hongfang; Di Xiaoyun; Chang Jinjia; Shen Youqing; Fan Weimin

    2012-01-01

    Background and purpose: We previously demonstrated that radiation may arrest tumor cells at G2 phase, which in turn prevents the cytotoxicity of antimicrotubule drugs and results in antagonistic interaction between these two modalities. Herein we tested whether G2 abrogators would attenuate the above antagonistic interaction and improve the therapeutic efficacy of combination therapy between radiation and antimicrotubule drugs. Materials and methods: Breast cancer BCap37 and epidermoid carcinoma KB cell lines were administered with radiation, UCN-01 (a model drug of G2 abrogator), paclitaxel or vincristine, alone or in combinations. The antitumor activities of single and combined treatments were analyzed by a series of cytotoxic, apoptotic, cell cycle, morphological and biochemical assays. Results: UCN-01 significantly enhanced the cytotoxicity of radiation, antimitotic drugs, and their combined treatments in vitro. Further investigations demonstrated that UCN-01 attenuated radiation-induced G2 arrest, and subsequently repressed the inhibitory effect of radiation on drug-induced mitotic arrest and apoptosis. Conclusions: This is the first report demonstrating that G2 checkpoint abrogation represses the inhibitory effect of radiation on antimicrotubule drugs, which may be implicated in cancer combination therapy. Considering that G2 abrogators are under extensive evaluation for cancer treatment, our findings provide valuable information for this class of promising compounds.

  6. ATM and KAT5 safeguard replicating chromatin against formaldehyde damage

    Science.gov (United States)

    Ortega-Atienza, Sara; Wong, Victor C.; DeLoughery, Zachary; Luczak, Michal W.; Zhitkovich, Anatoly

    2016-01-01

    Many carcinogens damage both DNA and protein constituents of chromatin, and it is unclear how cells respond to this compound injury. We examined activation of the main DNA damage-responsive kinase ATM and formation of DNA double-strand breaks (DSB) by formaldehyde (FA) that forms histone adducts and replication-blocking DNA-protein crosslinks (DPC). We found that low FA doses caused a strong and rapid activation of ATM signaling in human cells, which was ATR-independent and restricted to S-phase. High FA doses inactivated ATM via its covalent dimerization and formation of larger crosslinks. FA-induced ATM signaling showed higher CHK2 phosphorylation but much lower phospho-KAP1 relative to DSB inducers. Replication blockage by DPC did not produce damaged forks or detectable amounts of DSB during the main wave of ATM activation, which did not require MRE11. Chromatin-monitoring KAT5 (Tip60) acetyltransferase was responsible for acetylation and activation of ATM by FA. KAT5 and ATM were equally important for triggering of intra-S-phase checkpoint and ATM signaling promoted recovery of normal human cells after low-dose FA. Our results revealed a major role of the KAT5-ATM axis in protection of replicating chromatin against damage by the endogenous carcinogen FA. PMID:26420831

  7. Pyruvate Kinase M2 Is Required for the Expression of the Immune Checkpoint PD-L1 in Immune Cells and Tumors

    Directory of Open Access Journals (Sweden)

    Eva M. Palsson-McDermott

    2017-10-01

    Full Text Available Blocking interaction of the immune checkpoint receptor PD-1 with its ligand PD-L1 is associated with good clinical outcomes in a broad variety of malignancies. High levels of PD-L1 promote tumor growth by restraining CD8+ T-cell responses against tumors. Limiting PD-L1 expression and function is therefore critical for allowing the development of antitumor immune responses and effective tumor clearance. Pyruvate kinase isoform M2 (PKM2 is also a key player in regulating cancer as well as immune responses. PKM2 catalyzes the final rate-limiting step of glycolysis. Furthermore, PKM2 as a dimer translocates to the nucleus, where it stimulates hypoxia-inducible factor 1α (Hif-1α transactivation domain function and recruitment of p300 to the hypoxia response elements (HRE of Hif-1α target genes. Here, we provide the first evidence of a role for PKM2 in regulating the expression of PD-L1 on macrophages, dendritic cells (DCs, T cells, and tumor cells. LPS-induced expression of PD-L1 in primary macrophages was inhibited by the PKM2 targeting compound TEPP-46. Furthermore, RNA silencing of PKM2 inhibited LPS-induced PD-L1 expression. This regulation occurs through direct binding of PKM2 and Hif-1α to HRE sites on the PD-L1 promoter. Moreover, TEPP-46 inhibited expression of PD-L1 on macrophages, DCs, and T cells as well as tumor cells in a mouse CT26 cancer model. These findings broaden our understanding of how PKM2 may contribute to tumor progression and may explain the upregulation of PD-L1 in the tumor microenvironment.

  8. Coupling mechanisms between nucleosome assembly and the cellular response to DNA damage

    International Nuclear Information System (INIS)

    Lautrette, Aurelie

    2006-01-01

    Cells are continuously exposed to genotoxic stresses that induce a variety of DNA lesions. To protect their genome, cells have specific pathways that orchestrate the detection, signaling and repair of DNA damages. This work is dedicated to the characterization of such pathways that couple the DNA damage response to the assembly of chromatin, a complex that protects and regulates DNA accessibility. We have focused our study on two multifunctional proteins: Rad53, a central checkpoint kinase in the cellular response to DNA damage and Asf1, a histone chaperone involved in chromatin assembly. We have characterized in vitro the binding mode of Asf1 with Rad53 and Asfl with histones. This study is associated with the functional analysis of the role of these interactions in vivo in yeast cells. (author) [fr

  9. Replication stress, DNA damage signalling, and cytomegalovirus infection in human medulloblastomas

    DEFF Research Database (Denmark)

    Bartek, Jiri; Fornara, Olesja; Merchut-Maya, Joanna Maria

    2017-01-01

    suppressor activation, across our medulloblastoma cohort. Most tumours showed high proliferation (Ki67 marker), variable oxidative DNA damage (8-oxoguanine lesions) and formation of 53BP1 nuclear 'bodies', the latter indicating (along with ATR-Chk1 signalling) endogenous replication stress. The bulk...... cell replication stress and DNA repair. Collectively, the scenario we report here likely fuels genomic instability and evolution of medulloblastoma resistance to standard-of-care genotoxic treatments....... eight established immunohistochemical markers to assess the status of the DDR machinery, we found pronounced endogenous DNA damage signalling (γH2AX marker) and robust constitutive activation of both the ATM-Chk2 and ATR-Chk1 DNA damage checkpoint kinase cascades, yet unexpectedly modest p53 tumour...

  10. Dpb11/TopBP1 contributes to genomicstability via homologous recombinationand checkpoint signaling

    DEFF Research Database (Denmark)

    Germann, Susanne Manuela

    for recruitment. Also, the chicken homologue TopBP1 colocalizes with RPA1 as well as Rad51 when DNA damage is induced. Previously, dpb11 mutants have been shown to be sensitive to DNA-damaging agents that cause DSBs, DNA alkylation and stalled replication forks. Interestingly, we found the point mutants dpb11-PF...

  11. Damage analysis: damage function development and application

    International Nuclear Information System (INIS)

    Simons, R.L.; Odette, G.R.

    1975-01-01

    The derivation and application of damage functions, including recent developments for the U.S. LMFBR and CTR programs, is reviewed. A primary application of damage functions is in predicting component life expectancies; i.e., the fluence required in a service spectrum to attain a specified design property change. An important part of the analysis is the estimation of the uncertainty in such fluence limit predictions. The status of standardizing the procedures for the derivation and application of damage functions is discussed. Improvements in several areas of damage function development are needed before standardization can be completed. These include increasing the quantity and quality of the data used in the analysis, determining the limitations of the analysis due to the presence of multiple damage mechanisms, and finally, testing of damage function predictions against data obtained from material surveillance programs in operating thermal and fast reactors. 23 references. (auth)

  12. Spinal cord stimulation

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/007560.htm Spinal cord stimulation To use the sharing features on this page, please enable JavaScript. Spinal cord stimulation is a treatment for pain that uses ...

  13. Feldspar, Infrared Stimulated Luminescence

    DEFF Research Database (Denmark)

    Jain, Mayank

    2014-01-01

    This entry primarily concerns the characteristics and the origins of infrared-stimulated luminescence in feldspars.......This entry primarily concerns the characteristics and the origins of infrared-stimulated luminescence in feldspars....

  14. Growth hormone stimulation test

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003377.htm Growth hormone stimulation test To use the sharing features on this page, please enable JavaScript. The growth hormone (GH) stimulation test measures the ability of ...

  15. TNF-alpha impairs the S-G2/M cell cycle checkpoint and cyclobutane pyrimidine dimer repair in premalignant skin cells: Role of the PI3K-Akt pathway

    DEFF Research Database (Denmark)

    Faurschou, A.; Gniadecki, R.; Calay, D.

    2008-01-01

    Tumor necrosis factor-alpha (TNF-alpha) is induced by UVB radiation and has been implicated in the early stages of skin carcinogenesis. Here, we show that in normal keratinocytes and the transformed keratinocyte cell lines, HaCaT and A431, TNF-alpha stimulates protein kinase B/Akt, which results...... cycling. TNF-alpha enhanced apoptosis less potently and did not increase the level of CPD or stimulate cell cycle progression in normal keratinocytes. Our data suggest that TNF-alpha overrides the G2/M checkpoint in premalignant skin cells and allows for some cells containing unrepaired CPD to enter...... in activation of the survival complex mTORC1 (mammalian target of rapamycin complex 1) and inhibition of the proapoptotic proteins Bad and Fox03a. In UVB-irradiated HaCaT cells (10-20 mJ cm(-2)), TNF-alpha increased the proportion of cycling cells and enhanced the rate of apoptosis. A significantly higher...

  16. The Feasibility and Safety of Surgery in Patients Receiving Immune Checkpoint Inhibitors: A Retrospective Study

    Directory of Open Access Journals (Sweden)

    Alexandra W. Elias

    2017-06-01

    Full Text Available Immune checkpoint inhibitors (ICI are revolutionizing care for cancer patients. The list of malignancies for which the Food and Drug Administration is granting approval is rapidly increasing. Furthermore, there is a concomitant increase in clinical trials incorporating ICI. However, the safety of ICI in patients undergoing surgery remains unclear. Herein, we assessed the safety of ICI in the perioperative setting at a single center. We conducted a retrospective review of patients who underwent planned surgery while receiving ICI in the perioperative setting from 2012 to 2016. We collected 30-day postoperative morbidity and mortality utilizing the Clavien–Dindo classification system. We identified 17 patients who received perioperative ICI in 22 operations. Patients were diagnosed with melanoma (n = 14, renal cell carcinoma (n = 2, and urothelial carcinoma (n = 1. Therapies included pembrolizumab (n = 10, ipilimumab (n = 5, atezolizumab (n = 5, and ipilimumab/nivolumab (n = 2. Procedures included cutaneous/subcutaneous resection (n = 6, lymph node resection (n = 5, small bowel resection (n = 5, abdominal wall resection (n = 3, other abdominal surgery (n = 3, orthopedic surgery (n = 1, hepatic resection (n = 1, and neurosurgery (n = 2. There were no Grade III–IV Clavien–Dindo complications. There was one death secondary to ventricular fibrillation in the setting of coronary artery disease. ICI appear safe in the perioperative setting, involving multiple different types of surgery, and likely do not need to be stopped in the perioperative setting. Further studies are warranted to confirm these findings.

  17. Regulation of cell cycle checkpoint kinase WEE1 by miR-195 in malignant melanoma.

    Science.gov (United States)

    Bhattacharya, A; Schmitz, U; Wolkenhauer, O; Schönherr, M; Raatz, Y; Kunz, M

    2013-06-27

    WEE1 kinase has been described as a major gate keeper at the G2 cell cycle checkpoint and to be involved in tumour progression in different malignant tumours. Here we analysed the expression levels of WEE1 in a series of melanoma patient samples and melanoma cell lines using immunoblotting, quantitative real-time PCR and immunohistochemistry. WEE1 expression was significantly downregulated in patient samples of metastatic origin as compared with primary melanomas and in melanoma cell lines of high aggressiveness as compared with cell lines of low aggressiveness. Moreover, there was an inverse correlation between the expression of WEE1 and WEE1-targeting microRNA miR-195. Further analyses showed that transfection of melanoma cell lines with miR-195 indeed reduced WEE1 mRNA and protein expression in these cells. Reporter gene analysis confirmed direct targeting of the WEE1 3' untranslated region (3'UTR) by miR-195. Overexpression of miR-195 in SK-Mel-28 melanoma cells was accompanied by WEE1 reduction and significantly reduced stress-induced G2-M cell cycle arrest, which could be restored by stable overexpression of WEE1. Moreover, miR-195 overexpression and WEE1 knockdown, respectively, increased melanoma cell proliferation. miR-195 overexpression also enhanced migration and invasiveness of melanoma cells. Taken together, the present study shows that WEE1 expression in malignant melanoma is directly regulated by miR-195. miR-195-mediated downregulation of WEE1 in metastatic lesions may help to overcome cell cycle arrest under stress conditions in the local tissue microenvironment to allow unrestricted growth of tumour cells.

  18. How oocytes try to get it right: spindle checkpoint control in meiosis.

    Science.gov (United States)

    Touati, Sandra A; Wassmann, Katja

    2016-06-01

    The generation of a viable, diploid organism depends on the formation of haploid gametes, oocytes, and spermatocytes, with the correct number of chromosomes. Halving the genome requires the execution of two consecutive specialized cell divisions named meiosis I and II. Unfortunately, and in contrast to male meiosis, chromosome segregation in oocytes is error prone, with human oocytes being extraordinarily "meiotically challenged". Aneuploid oocytes, that are with the wrong number of chromosomes, give rise to aneuploid embryos when fertilized. In humans, most aneuploidies are lethal and result in spontaneous abortions. However, some trisomies survive to birth or even adulthood, such as the well-known trisomy 21, which gives rise to Down syndrome (Nagaoka et al. in Nat Rev Genet 13:493-504, 2012). A staggering 20-25 % of oocytes ready to be fertilized are aneuploid in humans. If this were not bad enough, there is an additional increase in meiotic missegregations as women get closer to menopause. A woman above 40 has a risk of more than 30 % of getting pregnant with a trisomic child. Worse still, in industrialized western societies, child birth is delayed, with women getting their first child later in life than ever. This trend has led to an increase of trisomic pregnancies by 70 % in the last 30 years (Nagaoka et al. in Nat Rev Genet 13:493-504, 2012; Schmidt et al. in Hum Reprod Update 18:29-43, 2012). To understand why errors occur so frequently during the meiotic divisions in oocytes, we review here the molecular mechanisms at works to control chromosome segregation during meiosis. An important mitotic control mechanism, namely the spindle assembly checkpoint or SAC, has been adapted to the special requirements of the meiotic divisions, and this review will focus on our current knowledge of SAC control in mammalian oocytes. Knowledge on how chromosome segregation is controlled in mammalian oocytes may help to identify risk factors important for questions

  19. Differential immune microenvironments and response to immune checkpoint blockade amongst molecular subtypes of murine medulloblastoma

    Science.gov (United States)

    Pham, Christina D.; Flores, Catherine; Yang, Changlin; Pinheiro, Elaine M.; Yearley, Jennifer H.; Sayour, Elias J.; Pei, Yanxin; Moore, Colin; McLendon, Roger E.; Huang, Jianping; Sampson, John H.; Wechsler-Reya, Robert; Mitchell, Duane A.

    2016-01-01

    PURPOSE Despite significant strides in the identification and characterization of potential therapeutic targets for medulloblastoma (MB), the role of the immune system and its interplay with the tumor microenvironment within these tumors are poorly understood. To address this, we adapted two syngeneic animal models of human Sonic Hedgehog (SHH)-driven and Group 3 MB for preclinical evaluation in immunocompetent C57BL/6 mice. METHODS AND RESULTS Multicolor flow cytometric analyses were used to phenotype and characterize immune infiltrating cells within established cerebellar tumors. We observed significantly higher percentages of dendritic cells, infiltrating lymphocytes, myeloid derived suppressor cells and tumor-associated macrophages in murine SHH model tumors compared with Group 3 tumors. However, murine Group 3 tumors had higher percentages of CD8+ PD-1+ T cells within the CD3 population. PD-1 blockade conferred superior antitumor efficacy in animals bearing intracranial Group 3 tumors compared to SHH group tumors, indicating that immunologic differences within the tumor microenvironment can be leveraged as potential targets to mediate antitumor efficacy. Further analysis of anti-PD-1 monoclonal antibody localization revealed binding to PD-1+ peripheral T cells, but not tumor infiltrating lymphocytes within the brain tumor microenvironment. Peripheral PD-1 blockade additionally resulted in a marked increase in CD3+ T cells within the tumor microenvironment. CONCLUSIONS This is the first immunologic characterization of preclinical models of molecular subtypes of MB and demonstration that response to immune checkpoint blockade differs across subtype classification. Our findings also suggest that effective anti-PD-1 blockade does not require that systemically administered antibodies penetrate the brain tumor microenvironment. PMID:26405194

  20. MK3 modulation affects BMI1-dependent and independent cell cycle check-points.

    Directory of Open Access Journals (Sweden)

    Peggy Prickaerts

    Full Text Available Although the MK3 gene was originally found deleted in some cancers, it is highly expressed in others. The relevance of MK3 for oncogenesis is currently not clear. We recently reported that MK3 controls ERK activity via a negative feedback mechanism. This prompted us to investigate a potential role for MK3 in cell proliferation. We here show that overexpression of MK3 induces a proliferative arrest in normal diploid human fibroblasts, characterized by enhanced expression of replication stress- and senescence-associated markers. Surprisingly, MK3 depletion evokes similar senescence characteristics in the fibroblast model. We previously identified MK3 as a binding partner of Polycomb Repressive Complex 1 (PRC1 proteins. In the current study we show that MK3 overexpression results in reduced cellular EZH2 levels and concomitant loss of epigenetic H3K27me3-marking and PRC1/chromatin-occupation at the CDKN2A/INK4A locus. In agreement with this, the PRC1 oncoprotein BMI1, but not the PCR2 protein EZH2, bypasses MK3-induced senescence in fibroblasts and suppresses P16INK4A expression. In contrast, BMI1 does not rescue the MK3 loss-of-function phenotype, suggesting the involvement of multiple different checkpoints in gain and loss of MK3 function. Notably, MK3 ablation enhances proliferation in two different cancer cells. Finally, the fibroblast model was used to evaluate the effect of potential tumorigenic MK3 driver-mutations on cell proliferation and M/SAPK signaling imbalance. Taken together, our findings support a role for MK3 in control of proliferation and replicative life-span, in part through concerted action with BMI1, and suggest that the effect of MK3 modulation or mutation on M/SAPK signaling and, ultimately, proliferation, is cell context-dependent.

  1. Differential Immune Microenvironments and Response to Immune Checkpoint Blockade among Molecular Subtypes of Murine Medulloblastoma.

    Science.gov (United States)

    Pham, Christina D; Flores, Catherine; Yang, Changlin; Pinheiro, Elaine M; Yearley, Jennifer H; Sayour, Elias J; Pei, Yanxin; Moore, Colin; McLendon, Roger E; Huang, Jianping; Sampson, John H; Wechsler-Reya, Robert; Mitchell, Duane A

    2016-02-01

    Despite significant strides in the identification and characterization of potential therapeutic targets for medulloblastoma, the role of the immune system and its interplay with the tumor microenvironment within these tumors are poorly understood. To address this, we adapted two syngeneic animal models of human Sonic Hedgehog (SHH)-driven and group 3 medulloblastoma for preclinical evaluation in immunocompetent C57BL/6 mice. Multicolor flow cytometric analyses were used to phenotype and characterize immune infiltrating cells within established cerebellar tumors. We observed significantly higher percentages of dendritic cells, infiltrating lymphocytes, myeloid-derived suppressor cells, and tumor-associated macrophages in murine SHH model tumors compared with group 3 tumors. However, murine group 3 tumors had higher percentages of CD8(+) PD-1(+) T cells within the CD3 population. PD-1 blockade conferred superior antitumor efficacy in animals bearing intracranial group 3 tumors compared with SHH group tumors, indicating that immunologic differences within the tumor microenvironment can be leveraged as potential targets to mediate antitumor efficacy. Further analysis of anti-PD-1 monoclonal antibody localization revealed binding to PD-1(+) peripheral T cells, but not tumor infiltrating lymphocytes within the brain tumor microenvironment. Peripheral PD-1 blockade additionally resulted in a marked increase in CD3(+) T cells within the tumor microenvironment. This is the first immunologic characterization of preclinical models of molecular subtypes of medulloblastoma and demonstration that response to immune checkpoint blockade differs across subtype classification. Our findings also suggest that effective anti-PD-1 blockade does not require that systemically administered antibodies penetrate the brain tumor microenvironment. ©2015 American Association for Cancer Research.

  2. Indoor Smartphone Navigation Using a Combination of Wi-Fi and Inertial Navigation with Intelligent Checkpoints

    Science.gov (United States)

    Hofer, H.; Retscher, G.

    2017-09-01

    For Wi-Fi positioning location fingerprinting is one of the most commonly employed localization technique. To achieve an acceptable level of positioning accuracy on the few meter level, i.e., to provide at least room resolution in buildings, such an approach is very labour consuming as it requires a high density of reference points. Thus, the novel approach developed aims at a significant reduction of workload for the training phase. The basic idea is to intelligently choose waypoints along possible users' trajectories in the indoor environment. These waypoints are termed intelligent checkpoints (iCPs) and serve as reference points for the fingerprinting localization approach. They are selected along the trajectories in such a way that they define a logical sequence with their ascending order. Thereby, the iCPs are located, for instance, at doors at entrances to buildings, rooms, along corridors, etc., or in low density along the trajectory to provide a suitable absolute user localization. Continuous positioning between these iCPs is obtained with the help of the smartphones' inertial sensors. While walking along a selected trajectory to the destination a dynamic recognition of the iCPs is performed and the drift of the inertial sensors is reduced as the iCP recognition serves as absolute position update. Conducted experiments in a multi-storey office building have shown that positioning accuracy of around 2.0 m are achievable which goes along with a reduction of workload by three quarter using this novel approach. The iCP concept and performance are presented and demonstrated in this paper.

  3. DNA-Damage Response RNA-Binding Proteins (DDRBPs): Perspectives from a New Class of Proteins and Their RNA Targets.

    Science.gov (United States)

    Dutertre, Martin; Vagner, Stéphan

    2017-10-27

    Upon DNA damage, cells trigger an early DNA-damage response (DDR) involving DNA repair and cell cycle checkpoints, and late responses involving gene expression regulation that determine cell fate. Screens for genes involved in the DDR have found many RNA-binding proteins (RBPs), while screens for novel RBPs have identified DDR proteins. An increasing number of RBPs are involved in early and/or late DDR. We propose to call this new class of actors of the DDR, which contain an RNA-binding activity, DNA-damage response RNA-binding proteins (DDRBPs). We then discuss how DDRBPs contribute not only to gene expression regulation in the late DDR but also to early DDR signaling, DNA repair, and chromatin modifications at DNA-damage sites through interactions with both long and short noncoding RNAs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Squalene Inhibits ATM-Dependent Signaling in γIR-Induced DNA Damage Response through Induction of Wip1 Phosphatase.

    Directory of Open Access Journals (Sweden)

    Naoto Tatewaki

    Full Text Available Ataxia telangiectasia mutated (ATM kinase plays a crucial role as a master controller in the cellular DNA damage response. Inhibition of ATM leads to inhibition of the checkpoint signaling pathway. Hence, addition of checkpoint inhibitors to anticancer therapies may be an effective targeting strategy. A recent study reported that Wip1, a protein phosphatase, de-phosphorylates serine 1981 of ATM during the DNA damage response. Squalene has been proposed to complement anticancer therapies such as chemotherapy and radiotherapy; however, there is little mechanistic information supporting this idea. Here, we report the inhibitory effect of squalene on ATM-dependent DNA damage signals. Squalene itself did not affect cell viability and the cell cycle of A549 cells, but it enhanced the cytotoxicity of gamma-irradiation (γIR. The in vitro kinase activity of ATM was not altered by squalene. However, squalene increased Wip1 expression in cells and suppressed ATM activation in γIR-treated cells. Consistent with the potential inhibition of ATM by squalene, IR-induced phosphorylation of ATM effectors such as p53 (Ser15 and Chk1 (Ser317 was inhibited by cell treatment with squalene. Thus, squalene inhibits the ATM-dependent signaling pathway following DNA damage through intracellular induction of Wip1 expression.

  5. DNA repair in PHA stimulated human lymphocytes

    International Nuclear Information System (INIS)

    Catena, C.; Mattoni, A.

    1984-01-01

    Damage an repair of radiation induced DNA strand breaks were measured by alkaline lysis and hydroxyapatite chromatography. PHA stimulated human lymphocytes show that the rejoining process is complete within the first 50 min., afterwords secondary DNA damage and chromatid aberration. DNA repair, in synchronized culture, allows to evaluate individual repair capacity and this in turn can contribute to the discovery of individual who, although they do not demonstrate apparent clinical signs, are carriers of DNA repair deficiency. Being evident that a correlation exists between DNA repair capacity and carcinogenesis, the possibility of evaluating the existent relationship between DNA repair and survival in tumor cells comes therefore into discussion

  6. Radiation damage of nonmetallic solids

    International Nuclear Information System (INIS)

    Goland, A.N.

    1975-01-01

    A review of data and information on radiation damage in nonmetallic solids is presented. Discussions are included on defects in nonmetals, radiation damage processes in nonmetals, electronic damage processes, physical damage processes, atomic displacement, photochemical damage processes, and ion implantation

  7. Maintaining Genome Stability in Defiance of Mitotic DNA Damage

    Science.gov (United States)

    Ferrari, Stefano; Gentili, Christian

    2016-01-01

    The implementation of decisions affecting cell viability and proliferation is based on prompt detection of the issue to be addressed, formulation and transmission of a correct set of instructions and fidelity in the execution of orders. While the first and the last are purely mechanical processes relying on the faithful functioning of single proteins or macromolecular complexes (sensors and effectors), information is the real cue, with signal amplitude, duration, and frequency ultimately determining the type of response. The cellular response to DNA damage is no exception to the rule. In this review article we focus on DNA damage responses in G2 and Mitosis. First, we set the stage describing mitosis and the machineries in charge of assembling the apparatus responsible for chromosome alignment and segregation as well as the inputs that control its function (checkpoints). Next, we examine the type of issues that a cell approaching mitosis might face, presenting the impact of post-translational modifications (PTMs) on the correct and timely functioning of pathways correcting errors or damage before chromosome segregation. We conclude this essay with a perspective on the current status of mitotic signaling pathway inhibitors and their potential use in cancer therapy. PMID:27493659

  8. Vaccination targeting human HER3 alters the phenotype of infiltrating T cells and responses to immune checkpoint inhibition.

    Science.gov (United States)

    Osada, Takuya; Morse, Michael A; Hobeika, Amy; Diniz, Marcio A; Gwin, William R; Hartman, Zachary; Wei, Junping; Guo, Hongtao; Yang, Xiao-Yi; Liu, Cong-Xiao; Kaneko, Kensuke; Broadwater, Gloria; Lyerly, H Kim

    2017-01-01

    Expression of human epidermal growth factor family member 3 (HER3), a critical heterodimerization partner with EGFR and HER2, promotes more aggressive biology in breast and other epithelial malignancies. As such, inhibiting HER3 could have broad applicability to the treatment of EGFR- and HER2-driven tumors. Although lack of a functional kinase domain limits the use of receptor tyrosine kinase inhibitors, HER3 contains antigenic targets for T cells and antibodies. Using novel human HER3 transgenic mouse models of breast cancer, we demonstrate that immunization with recombinant adenoviral vectors encoding full length human HER3 (Ad-HER3-FL) induces HER3-specific T cells and antibodies, alters the T cell infiltrate in tumors, and influences responses to immune checkpoint inhibitions. Both preventative and therapeutic Ad-HER3-FL immunization delayed tumor growth but were associated with both intratumoral PD-1 expressing CD8 + T cells and regulatory CD4 + T cell infiltrates. Immune checkpoint inhibition with either anti-PD-1 or anti-PD-L1 antibodies increased intratumoral CD8 + T cell infiltration and eliminated tumor following preventive vaccination with Ad-HER3-FL vaccine. The combination of dual PD-1/PD-L1 and CTLA4 blockade slowed the growth of tumor in response to Ad-HER3-FL in the therapeutic model. We conclude that HER3-targeting vaccines activate HER3-specific T cells and induce anti-HER3 specific antibodies, which alters the intratumoral T cell infiltrate and responses to immune checkpoint inhibition.

  9. Pch2 acts through Xrs2 and Tel1/ATM to modulate interhomolog bias and checkpoint function during meiosis.

    Directory of Open Access Journals (Sweden)

    Hsuan-Chung Ho

    2011-11-01

    Full Text Available Proper segregation of chromosomes during meiosis requires the formation and repair of double-strand breaks (DSBs to form crossovers. Repair is biased toward using the homolog as a substrate rather than the sister chromatid. Pch2 is a conserved member of the AAA(+-ATPase family of proteins and is implicated in a wide range of meiosis-specific processes including the recombination checkpoint, maturation of the chromosome axis, crossover control, and synapsis. We demonstrate a role for Pch2 in promoting and regulating interhomolog bias and the meiotic recombination checkpoint in response to unprocessed DSBs through the activation of axial proteins Hop1 and Mek1 in budding yeast. We show that Pch2 physically interacts with the putative BRCT repeats in the N-terminal region of Xrs2, a member of the MRX complex that acts at sites of unprocessed DSBs. Pch2, Xrs2, and the ATM ortholog Tel1 function in the same pathway leading to the phosphorylation of Hop1, independent of Rad17 and the ATR ortholog Mec1, which respond to the presence of single-stranded DNA. An N-terminal deletion of Xrs2 recapitulates the pch2Δ phenotypes for signaling unresected breaks. We propose that interaction with Xrs2 may enable Pch2 to remodel chromosome structure adjacent to the site of a DSB and thereby promote accessibility of Hop1 to the Tel1 kinase. In addition, Xrs2, like Pch2, is required for checkpoint-mediated delay conferred by the failure to synapse chromosomes.

  10. Immune Checkpoint Molecules on Tumor-Infiltrating Lymphocytes and Their Association with Tertiary Lymphoid Structures in Human Breast Cancer

    Directory of Open Access Journals (Sweden)

    Cinzia Solinas

    2017-10-01

    Full Text Available There is an exponentially growing interest in targeting immune checkpoint molecules in breast cancer (BC, particularly in the triple-negative subtype where unmet treatment needs remain. This study was designed to analyze the expression, localization, and prognostic role of PD-1, PD-L1, PD-L2, CTLA-4, LAG3, and TIM3 in primary BC. Gene expression analysis using the METABRIC microarray dataset found that all six immune checkpoint molecules are highly expressed in basal-like and HER2-enriched compared to the other BC molecular subtypes. Flow cytometric analysis of fresh tissue homogenates from untreated primary tumors show that PD-1 is principally expressed on CD4+ or CD8+ T cells and CTLA-4 is expressed on CD4+ T cells. The global proportion of PD-L1+, PD-L2+, LAG3+, and TIM3+ tumor-infiltrating lymphocytes (TIL was low and detectable in only a small number of tumors. Immunohistochemically staining fixed tissues from the same tumors was employed to score TIL and tertiary lymphoid structures (TLS. PD-L1+, PD-L2+, LAG3+, and TIM3+ cells were detected in some TLS in a pattern that resembles secondary lymphoid organs. This observation suggests that TLS are important sites of immune activation and regulation, particularly in tumors with extensive baseline immune infiltration. Significantly improved overall survival was correlated with PD-1 expression in the HER2-enriched and PD-L1 or CTLA-4 expression in basal-like BC. PD-1 and CTLA-4 proteins were most frequently detected on TIL, which supports the correlations observed between their gene expression and improved long-term outcome in basal-like and HER2-enriched BC. PD-L1 expression by tumor or immune cells is uncommon in BC. Overall, the data presented here distinguish PD-1 as a marker of T cell activity in both the T and B cell areas of BC associated TLS. We found that immune checkpoint molecule expression parallels the extent of TIL and TLS, although there is a noteworthy amount of heterogeneity

  11. DNA damage by X-rays and their impact on replication processes

    International Nuclear Information System (INIS)

    Parplys, Ann Christin; Petermann, Eva; Petersen, Cordula; Dikomey, Ekkehard; Borgmann, Kerstin

    2012-01-01

    Background: Replication-dependent radiosensitization of tumors ranks among the most promising tools for future improvements in tumor therapy. However, cell cycle checkpoint signaling during S phase is a key for maintaining genomic stability after ionizing irradiation allowing DNA damage repair by stabilizing replication forks, inhibiting new origin firing and recruiting DNA repair proteins. As the impact of the different types of DNA damage induced by ionizing radiation on replication fork functionality has not been investigated, this study was performed in tumor cells treated with various agents that induce specific DNA lesions. Methods: U2OS cells were exposed to methyl methanesulfonate (MMS) to induce base damage, low or high concentrations of hydrogen peroxide for the induction of SSBs, Topotecan to induce DSBs at replication, Mitomycin C (MMC) to induce interstrand cross-links or ionizing irradiation to analyze all damages. Chk1 phosphorylation, origin firing and replication fork progression, and cell cycle distribution were analyzed. Results: In our system, the extent of Chk1 phosphorylation was dependent on the type of damage induced and prolonged Chk1 phosphorylation correlated with the inhibition of replication initiation. Ionizing radiation, high concentrations of hydrogen peroxide, and Topotecan affected replication elongation much more strongly that the other agents. Almost all agents induced a slight increase in the S phase population but subsequent G2 arrest was only observed in response to those agents that strongly inhibited replication elongation and caused prolonged Chk1 phosphorylation. Conclusions: Our data suggest that to improve radiotherapy, radiosensitivity in S phase could be increased by combining irradiation with agents that induce secondary DSB or inhibit checkpoint signaling, such as inhibitors of PARP or Chk1.

  12. Binding of Multiple Rap1 Proteins Stimulates Chromosome Breakage Induction during DNA Replication.

    Directory of Open Access Journals (Sweden)

    Greicy H Goto

    2015-08-01

    Full Text Available Telomeres, the ends of linear eukaryotic chromosomes, have a specialized chromatin structure that provides a stable chromosomal terminus. In budding yeast Rap1 protein binds to telomeric TG repeat and negatively regulates telomere length. Here we show that binding of multiple Rap1 proteins stimulates DNA double-stranded break (DSB induction at both telomeric and non-telomeric regions. Consistent with the role of DSB induction, Rap1 stimulates nearby recombination events in a dosage-dependent manner. Rap1 recruits Rif1 and Rif2 to telomeres, but neither Rif1 nor Rif2 is required for DSB induction. Rap1-mediated DSB induction involves replication fork progression but inactivation of checkpoint kinase Mec1 does not affect DSB induction. Rap1 tethering shortens artificially elongated telomeres in parallel with telomerase inhibition, and this telomere shortening does not require homologous recombination. These results suggest that Rap1 contributes to telomere homeostasis by promoting chromosome breakage.

  13. Response of thyroid follicular cells to gamma irradiation compared to proton irradiation. I. Initial characterization of DNA damage, micronucleus formation, apoptosis, cell survival, and cell cycle phase redistribution

    Science.gov (United States)

    Green, L. M.; Murray, D. K.; Bant, A. M.; Kazarians, G.; Moyers, M. F.; Nelson, G. A.; Tran, D. T.

    2001-01-01

    The RBE of protons has been assumed to be equivalent to that of photons. The objective of this study was to determine whether radiation-induced DNA and chromosome damage, apoptosis, cell killing and cell cycling in organized epithelial cells was influenced by radiation quality. Thyroid-stimulating hormone-dependent Fischer rat thyroid cells, established as follicles, were exposed to gamma rays or proton beams delivered acutely over a range of physical doses. Gamma-irradiated cells were able to repair DNA damage relatively rapidly so that by 1 h postirradiation they had approximately 20% fewer exposed 3' ends than their counterparts that had been irradiated with proton beams. The persistence of free ends of DNA in the samples irradiated with the proton beam implies that either more initial breaks or a quantitatively different type of damage had occurred. These results were further supported by an increased frequency of chromosomal damage as measured by the presence of micronuclei. Proton-beam irradiation induced micronuclei at a rate of 2.4% per gray, which at 12 Gy translated to 40% more micronuclei than in comparable gamma-irradiated cultures. The higher rate of micronucleus formation and the presence of larger micronuclei in proton-irradiated cells was further evidence that a qualitatively more severe class of damage had been induced than was induced by gamma rays. Differences in the type of damage produced were detected in the apoptosis assay, wherein a significant lag in the induction of apoptosis occurred after gamma irradiation that did not occur with protons. The more immediate expression of apoptotic cells in the cultures irradiated with the proton beam suggests that the damage inflicted was more severe. Alternatively, the cell cycle checkpoint mechanisms required for recovery from such damage might not have been invoked. Differences based on radiation quality were also evident in the alpha components of cell survival curves (0.05 Gy(-1) for gamma rays, 0

  14. Femoral nerve damage (image)

    Science.gov (United States)

    The femoral nerve is located in the leg and supplies the muscles that assist help straighten the leg. It supplies sensation ... leg. One risk of damage to the femoral nerve is pelvic fracture. Symptoms of femoral nerve damage ...

  15. As we age: Does slippage of quality control in the immune system lead to collateral damage?

    Science.gov (United States)

    Müller, Ludmila; Pawelec, Graham

    2015-09-01

    The vertebrate adaptive immune system is remarkable for its possession of a very broad range of antigen receptors imbuing the system with exquisite specificity, in addition to the phagocytic and inflammatory cells of the innate system shared with invertebrates. This system requires strict control both at the level of the generation the cells carrying these receptors and at the level of their activation and effector function mediation in order to avoid autoimmunity and mitigate immune pathology. Thus, quality control checkpoints are built into the system at multiple nodes in the response, relying on clonal selection and regulatory networks to maximize pathogen-directed effects and minimize collateral tissue damage. However, these checkpoints are compromised with age, resulting in poorer immune control manifesting as tissue-damaging autoimmune and inflammatory phenomena which can cause widespread systemic disease, paradoxically compounding the problems associated with increased susceptibility to infectious disease and possibly cancer in the elderly. Better understanding the reasons for slippage of immune control will pave the way for developing rational strategies for interventions to maintain appropriate immunity while reducing immunopathology. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Mouse Rad9b is essential for embryonic development and promotes resistance to DNA damage

    Science.gov (United States)

    Leloup, Corinne; Hopkins, Kevin M.; Wang, Xiangyuan; Zhu, Aiping; Wolgemuth, Debra J.; Lieberman, Howard B.

    2010-01-01

    RAD9 participates in promoting resistance to DNA damage, cell cycle checkpoint control, DNA repair, apoptosis, embryogenesis, and regulation of transcription. A paralogue of RAD9 (named RAD9B) has been identified. To define the function of mouse Rad9b (Mrad9b), embryonic stem (ES) cells with a targeted gene deletion were constructed and used to generate Mrad9b mutant mice. Mrad9b−/− embryos are resorbed after E7.5 while some of the heterozygotes die between E12.5 and a few days after birth. Mrad9b is expressed in embryonic brain and Mrad9b+/− embryos exhibit abnormal neural tube closure. Mrad9b−/− mouse embryonic fibroblasts are not viable. Mrad9b−/− ES cells are more sensitive to gamma rays and mitomycin C than Mrad9b+/+ controls, but show normal gamma-ray-induced G2/M checkpoint control. There is no evidence of spontaneous genomic instability in Mrad9b−/− cells. Our findings thus indicate that Mrad9b is essential for embryonic development and mediates resistance to certain DNA damaging agents. PMID:20842695

  17. Checkpoint inhibitors in cancer immunotherapy: Cross reactivity of a CTLA-4 antibody and IDO-inhibitor L-1MT in pigs

    DEFF Research Database (Denmark)

    Al-Shatrawi, Zina Adil; Frøsig, Thomas Mørch; Jungersen, Gregers

    a non-specific activation of porcine T cells. This will be further investigated to provide the basis for in vivo studies investigating checkpoint inhibitor blockade in combination with other cancer immunotherapies. Eventually our goal is to establish pigs as an alternative large animal model......Blockade of checkpoint inhibitors has recently shown very convincing results in the treatment of cancer. One key target is CTLA-4, which has been demonstrated to be a potent negative regulator of lymphocyte activation. The treatment with the FDA-approved fully human CTLA-4 monoclonal antibody...... Ipilimumab increases anticancer T-cell reactivity and overall survival of metastatic cancer patients. Indole-amine 2,3-dioxygenase (IDO) is another checkpoint inhibitor which suppresses T-cell immunity by the depletion of tryptophan in the T-cell microenvironment, and also inhibition of IDO by L-1...

  18. Evolution of checkpoint inhibitors for the treatment of metastatic gastric cancers: Current status and future perspectives.

    Science.gov (United States)

    Taieb, Julien; Moehler, Markus; Boku, Narikazu; Ajani, Jaffer A; Yañez Ruiz, Eduardo; Ryu, Min-Hee; Guenther, Silke; Chand, Vikram; Bang, Yung-Jue

    2018-05-01

    Standard treatment options for patients with advanced gastric or gastroesophageal junction cancer (GC/GEJC) are associated with limited efficacy and some toxicity. Recently, immunotherapy with antibodies that inhibit the programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) interaction has emerged as a new treatment option. This manuscript reviews early-phase and late-phase trials of immunotherapy in advanced GC/GEJC. Searches for studies of immunotherapy in GC/GEJC were performed using PubMed, ClinicalTrials.gov, and abstract databases for select annual congresses. Findings were interpreted based on expert opinion. Monotherapy with anti-PD-1/PD-L1 antibodies, including pembrolizumab, nivolumab, avelumab, durvalumab, and atezolizumab, has shown interesting objective response rates (ORRs; 7-26%) across varying GC/GEJC populations, with ORRs potentially higher in PD-L1 + vs PD-L1 - tumors. Safety profiles compare favorably with chemotherapy, with grade ≥3 treatment-related adverse events occurring in 5-17%. Based on a large phase 2 study, pembrolizumab was approved in the United States for third-line treatment of patients with PD-L1 + GC/GEJC. In a phase 3 trial, third-line or later nivolumab increased overall survival vs placebo in an Asian population, leading to regulatory approval in Japan, although other completed phase 3 trials did not show superiority for pembrolizumab or avelumab monotherapy vs chemotherapy. Other trials in advanced GC/GEJC are assessing various anti-PD-1/PD-L1-based strategies, including administration in first-line and later-line settings and as combination (with chemotherapy or agents targeting other immune checkpoint proteins, eg, CTLA-4, LAG-3, and IDO) or switch-maintenance regimens. Anti-PD-1/PD-L1 antibodies have shown encouraging clinical activity in advanced GC/GEJC. Results from ongoing phase 3 trials are needed to further evaluate the potential roles of these agents within the continuum of care. Copyright

  19. The human papillomavirus type 58 E7 oncoprotein modulates cell cycle regulatory proteins and abrogates cell cycle checkpoints

    International Nuclear Information System (INIS)

    Zhang Weifang; Li Jing; Kanginakudru, Sriramana; Zhao Weiming; Yu Xiuping; Chen, Jason J.

    2010-01-01

    HPV type 58 (HPV-58) is the third most common HPV type in cervical cancer from Eastern Asia, yet little is known about how it promotes carcinogenesis. In this study, we demonstrate that HPV-58 E7 significantly promoted the proliferation and extended the lifespan of primary human keratinocytes (PHKs). HPV-58 E7 abrogated the G1 and the postmitotic checkpoints, although less efficiently than HPV-16 E7. Consistent with these observations, HPV-58 E7 down-regulated the cellular tumor suppressor pRb to a lesser extent than HPV-16 E7. Similar to HPV-16 E7 expressing PHKs, Cdk2 remained active in HPV-58 E7 expressing PHKs despite the presence of elevated levels of p53 and p21. Interestingly, HPV-58 E7 down-regulated p130 more efficiently than HPV-16 E7. Our study demonstrates a correlation between the ability of down-regulating pRb/p130 and abrogating cell cycle checkpoints by HPV-58 E7, which also correlates with the biological risks of cervical cancer progression associated with HPV-58 infection.

  20. Pemodelan Molekul Turunan p-Metoksi sinnamoil Hidrazida Sebagai Inhibitor Checkpoint Kinase 1 dan Inhibitor Aromatase secara In silico

    Directory of Open Access Journals (Sweden)

    Galih Satrio Putra

    2017-08-01

    Full Text Available The development of anticancer drugs from ethyl p-methoxycinnamate (EPMC derivatives continues to obtain compounds that have high ability of cancer cells apoptosis and minimal side effects. p-Methoxycinnamoyl hydrazide derivate compounds from EPMC structure modification were docked into the ligand-binding pocket of Check point kinase 1 enzymes (2YWP and the aromatase enzyme (3S7S using software Molegro Virtual Docker (MVD Ver.5.5. We compared the Rerank score of native ligand with derivate compounds of p-Methoxycinnamoyl hydrazide. Rerank scores of compounds 4b and 4c (-99.98 Kcal/mol and -99.80 Kcal/mol were lower than the native ligand A42 in inhibiting the enzyme checkpoint kinase 1. Rerank values of p-Methoxycinnamoyl hydrazide derivate compounds were greater than the native ligand EXM in inhibiting the enzyme aromatase. p-Methoxycinnamoyl hydrazide derivate compounds, especially compounds 4b and 4c, had anticancer mechanism by inhibiting the enzyme pathway checkpoint kinase 1 and had not activity in inhibiting the aromatase enzyme.

  1. Interactions of checkpoint-genes RAD9, RAD17, RAD24 and RAD53 determining radioresistance of Yeast Saccharomyces Cerevisiae

    International Nuclear Information System (INIS)

    Koltovaya, N.A.; Nikulushkina, Yu.V.; Roshchina, M.P.; Devin, A.B.

    2007-01-01

    The mechanisms of genetic control of progress through the division cell cycle (checkpoint-control) in yeast Saccharomyces cerevisiae have been studied intensively. To investigate the role of checkpoint-genes RAD9, RAD17, RAD24, RAD53 in cell radioresistance we have investigated cell sensitivity of double mutants to γ-ray. Double mutants involving various combinations with rad9Δ show epistatic interactions, i.e. the sensitivity of the double mutants to γ-ray was no greater than that of more sensitive of the two single mutants. This suggests that all these genes govern the same pathway. This group of genes was named RAD9-epistasis group. It is interesting to note that the genes RAD9 and RAD53 have positive effect but RAD17 and RAD24 have negative effect on radiosensitivity of yeast cells. Interactions between mutations may differ depending on the agent γ-ray or UV-light, for example mutations rad9Δ and rad24Δ show additive effect for γ-ray and epistatic effect for UV-light

  2. Cancer immunogenomic approach to neoantigen discovery in a checkpoint blockade responsive murine model of oral cavity squamous cell carcinoma

    Science.gov (United States)

    Zolkind, Paul; Przybylski, Dariusz; Marjanovic, Nemanja; Nguyen, Lan; Lin, Tianxiang; Johanns, Tanner; Alexandrov, Anton; Zhou, Liye; Allen, Clint T.; Miceli, Alexander P.; Schreiber, Robert D.; Artyomov, Maxim; Dunn, Gavin P.; Uppaluri, Ravindra

    2018-01-01

    Head and neck squamous cell carcinomas (HNSCC) are an ideal immunotherapy target due to their high mutation burden and frequent infiltration with lymphocytes. Preclinical models to investigate targeted and combination therapies as well as defining biomarkers to guide treatment represent an important need in the field. Immunogenomics approaches have illuminated the role of mutation-derived tumor neoantigens as potential biomarkers of response to checkpoint blockade as well as representing therapeutic vaccines. Here, we aimed to define a platform for checkpoint and other immunotherapy studies using syngeneic HNSCC cell line models (MOC2 and MOC22), and evaluated the association between mutation burden, predicted neoantigen landscape, infiltrating T cell populations and responsiveness of tumors to anti-PD1 therapy. We defined dramatic hematopoietic cell transcriptomic alterations in the MOC22 anti-PD1 responsive model in both tumor and draining lymph nodes. Using a cancer immunogenomics pipeline and validation with ELISPOT and tetramer analysis, we identified the H-2Kb-restricted ICAM1P315L (mICAM1) as a neoantigen in MOC22. Finally, we demonstrated that mICAM1 vaccination was able to protect against MOC22 tumor development defining mICAM1 as a bona fide neoantigen. Together these data define a pre-clinical HNSCC model system that provides a foundation for future investigations into combination and novel therapeutics. PMID:29423108

  3. Chromatin relaxation-mediated induction of p19INK4d increases the ability of cells to repair damaged DNA.

    Science.gov (United States)

    Ogara, María F; Sirkin, Pablo F; Carcagno, Abel L; Marazita, Mariela C; Sonzogni, Silvina V; Ceruti, Julieta M; Cánepa, Eduardo T

    2013-01-01

    The maintenance of genomic integrity is of main importance to the survival and health of organisms which are continuously exposed to genotoxic stress. Cells respond to DNA damage by activating survival pathways consisting of cell cycle checkpoints and repair mechanisms. However, the signal that triggers the DNA damage response is not necessarily a direct detection of the primary DNA lesion. In fact, chromatin defects may serve as initiating signals to activate those mechanisms. If the modulation of chromatin structure could initiate a checkpoint response in a direct manner, this supposes the existence of specific chromatin sensors. p19INK4d, a member of the INK4 cell cycle inhibitors, plays a crucial role in regulating genomic stability and cell viability by enhancing DNA repair. Its expression is induced in cells injured by one of several genotoxic treatments like cis-platin, UV light or neocarzinostatin. Nevertheless, when exogenous DNA damaged molecules are introduced into the cell, this induction is not observed. Here, we show that p19INK4d is enhanced after chromatin relaxation even in the absence of DNA damage. This induction was shown to depend upon ATM/ATR, Chk1/Chk2 and E2F activity, as is the case of p19INK4d induction by endogenous DNA damage. Interestingly, p19INK4d improves DNA repair when the genotoxic damage is caused in a relaxed-chromatin context. These results suggest that changes in chromatin structure, and not DNA damage itself, is the actual trigger of p19INK4d induction. We propose that, in addition to its role as a cell cycle inhibitor, p19INK4d could participate in a signaling network directed to detecting and eventually responding to chromatin anomalies.

  4. Chromatin relaxation-mediated induction of p19INK4d increases the ability of cells to repair damaged DNA.

    Directory of Open Access Journals (Sweden)

    María F Ogara

    Full Text Available The maintenance of genomic integrity is of main importance to the survival and health of organisms which are continuously exposed to genotoxic stress. Cells respond to DNA damage by activating survival pathways consisting of cell cycle checkpoints and repair mechanisms. However, the signal that triggers the DNA damage response is not necessarily a direct detection of the primary DNA lesion. In fact, chromatin defects may serve as initiating signals to activate those mechanisms. If the modulation of chromatin structure could initiate a checkpoint response in a direct manner, this supposes the existence of specific chromatin sensors. p19INK4d, a member of the INK4 cell cycle inhibitors, plays a crucial role in regulating genomic stability and cell viability by enhancing DNA repair. Its expression is induced in cells injured by one of several genotoxic treatments like cis-platin, UV light or neocarzinostatin. Nevertheless, when exogenous DNA damaged molecules are introduced into the cell, this induction is not observed. Here, we show that p19INK4d is enhanced after chromatin relaxation even in the absence of DNA damage. This induction was shown to depend upon ATM/ATR, Chk1/Chk2 and E2F activity, as is the case of p19INK4d induction by endogenous DNA damage. Interestingly, p19INK4d improves DNA repair when the genotoxic damage is caused in a relaxed-chromatin context. These results suggest that changes in chromatin structure, and not DNA damage itself, is the actual trigger of p19INK4d induction. We propose that, in addition to its role as a cell cycle inhibitor, p19INK4d could participate in a signaling network directed to detecting and eventually responding to chromatin anomalies.

  5. Ermenilerin Kontrol Noktası: İskenderun Limanı Alexandretta Port - Checkpoint For Armenians

    Directory of Open Access Journals (Sweden)

    Naim ÜRKMEZ

    2013-07-01

    Full Text Available The period of Yavuz Sultan Selim and the Ottoman Empire’s entering Alexandretta until the first half of the 19th century had identity of a very small village. Right to the end of the century, the industry revolution’s impact and trading goods’ large amount of transport means gained importance starting especially in the city of Aleppo and then in one of South Anatolia’s natural ports as of the day the preliminary plan came out. After the 1890’s the increase of Armenian activities in Anatolia and the outbreak of revolts in many other places parallel the Ottoman government’s large forces’ involvement to prevent and be able to keep Armenian rebels under control, taking various measures. Abdul Hamid II started under this negative initiative; the Sultan tried to resist in various ways. From these measures in any of the provinces of Aleppo, Mamuretülaziz, Adana, Kayseri, Bitlis, Van, Diyarbekir, or Erzurum, Armenians living in the diaspora and connecting supplier port, as in Alexandretta, have a checkpoint facility. Sultan Abdul Hamid II’s direction of the facility is seen at the checkpoint from the dock outside rising or Anatolia geography’s various residential areas incoming Armenian travelers often reserved monthly lists. In addition to this, England in the grip of Cyprus was able to check Armenians’ entries into Anatolia set by a weapons and bombs training field. It is understood that this matter is elaborated on further in books prepared from lists. These books will come to light with the acceleration of the classification of records. Yavuz Sultan Selim döneminde Osmanlı hâkimiyetine giren İskenderun, 19. yüzyılın ilk yarısına kadar çok küçük bir köy hüviyetinde kalmıştır. Yüzyılın sonuna doğru sanayi devriminin etkisi ve ticari emtianın çokça taşınması vesilesi ile önem kazanmaya başlayan şehir özellikle Halep ve ardı ile Güney Anadolu’nun doğal bir limanı olarak gün geçtikçe ön plana

  6. Phospho-Ser/Thr-binding domains: navigating the cell cycle and DNA damage response.

    Science.gov (United States)

    Reinhardt, H Christian; Yaffe, Michael B

    2013-09-01

    Coordinated progression through the cell cycle is a complex challenge for eukaryotic cells. Following genotoxic stress, diverse molecular signals must be integrated to establish checkpoints specific for each cell cycle stage, allowing time for various types of DNA repair. Phospho-Ser/Thr-binding domains have emerged as crucial regulators of cell cycle progression and DNA damage signalling. Such domains include 14-3-3 proteins, WW domains, Polo-box domains (in PLK1), WD40 repeats (including those in the E3 ligase SCF(βTrCP)), BRCT domains (including those in BRCA1) and FHA domains (such as in CHK2 and MDC1). Progress has been made in our understanding of the motif (or motifs) that these phospho-Ser/Thr-binding domains connect with on their targets and how these interactions influence the cell cycle and DNA damage response.

  7. ATR- and ATM-Mediated DNA Damage Response Is Dependent on Excision Repair Assembly during G1 but Not in S Phase of Cell Cycle.

    Science.gov (United States)

    Ray, Alo; Blevins, Chessica; Wani, Gulzar; Wani, Altaf A

    2016-01-01

    Cell cycle checkpoint is mediated by ATR and ATM kinases, as a prompt early response to a variety of DNA insults, and culminates in a highly orchestrated signal transduction cascade. Previously, we defined the regulatory role of nucleotide excision repair (NER) factors, DDB2 and XPC, in checkpoint and ATR/ATM-dependent repair pathway via ATR and ATM phosphorylation and recruitment to ultraviolet radiation (UVR)-induced damage sites. Here, we have dissected the molecular mechanisms of DDB2- and XPC- mediated regulation of ATR and ATM recruitment and activation upon UVR exposures. We show that the ATR and ATM activation and accumulation to UVR-induced damage not only depends on DDB2 and XPC, but also on the NER protein XPA, suggesting that the assembly of an active NER complex is essential for ATR and ATM recruitment. ATR and ATM localization and H2AX phosphorylation at the lesion sites occur as early as ten minutes in asynchronous as well as G1 arrested cells, showing that repair and checkpoint-mediated by ATR and ATM starts early upon UV irradiation. Moreover, our results demonstrated that ATR and ATM recruitment and H2AX phosphorylation are dependent on NER proteins in G1 phase, but not in S phase. We reasoned that in G1 the UVR-induced ssDNA gaps or processed ssDNA, and the bound NER complex promote ATR and ATM recruitment. In S phase, when the UV lesions result in stalled replication forks with long single-stranded DNA, ATR and ATM recruitment to these sites is regulated by different sets of proteins. Taken together, these results provide evidence that UVR-induced ATR and ATM recruitment and activation differ in G1 and S phases due to the existence of distinct types of DNA lesions, which promote assembly of different proteins involved in the process of DNA repair and checkpoint activation.

  8. Kin3 protein, a NIMA-related kinase of Saccharomyces cerevisiae, is involved in DNA adduct damage response.

    Science.gov (United States)

    Moura, Dinara J; Castilhos, Bruna; Immich, Bruna F; Cañedo, Andrés D; Henriques, João A P; Lenz, Guido; Saffi, Jenifer

    2010-06-01

    Kin3 is a nonessential serine/threonine protein kinase of the budding yeast Saccharomyces cerevisiae with unknown cellular role. It is an ortholog of the Aspergillus nidulans protein kinase NIMA (Never-In Mitosis, gene A), which is involved in the regulation of G2/M phase progression, DNA damage response and mitosis. The aim of this study was to determine whether Kin3 is required for proper checkpoint activation and DNA repair. Here we show that KIN3 gene deficient cells present sensitivity and fail to arrest properly at G2/M-phase checkpoint in response to the DNA damage inducing agents MMS, cisplatin, doxorubicin and nitrogen mustard, suggesting that Kin3 can be involved in DNA strand breaks recognition or signaling. In addition, there is an increase in KIN3 gene expression in response to the mutagenic treatment, which was confirmed by the increase of Kin3 protein. We also showed that co-treatment with caffeine induces a slight increase in the susceptibility to genotoxic agents in kin3 cells and abolishes KIN3 gene expression in wild-type strain, suggesting that Kin3p can play a role in Tel1/Mec1-dependent pathway activation induced after genotoxic stress. These data provide the first evidence of the involvement of S. cerevisiae Kin3 in the DNA damage response.

  9. Histone H2AX participates the DNA damage-induced ATM activation through interaction with NBS1.

    Science.gov (United States)

    Kobayashi, Junya; Tauchi, Hiroshi; Chen, Benjamin; Burma, Sandeep; Bruma, Sandeep; Tashiro, Satoshi; Matsuura, Shinya; Tanimoto, Keiji; Chen, David J; Komatsu, Kenshi

    2009-03-20

    Phosphorylated histone H2AX (gamma-H2AX) functions in the recruitment of DNA damage response proteins to DNA double-strand breaks (DSBs) and facilitates DSB repair. ATM also co-localizes with gamma-H2AX at DSB sites following its auto-phosphorylation. However, it is unclear whether gamma-H2AX has a role in activation of ATM-dependent cell cycle checkpoints. Here, we show that ATM as well as NBS1 is recruited to damaged-chromatin in a gamma-H2AX-dependent manner. Foci formation of phosphorylated ATM and ATM-dependent phosphorylation is repressed in H2AX-knockdown cells. Furthermore, anti-gamma-H2AX antibody co-immunoprecipitates an ATM-like protein kinase activity in vitro and recombinant H2AX increases in vitro kinase activity of ATM from un-irradiated cells. Moreover, H2AX-deficient cells exhibited a defect in ATM-dependent cell cycle checkpoints. Taken together, gamma-H2AX has important role for effective DSB-dependent activation of ATM-related damage responses via NBS1.

  10. Histone H2AX participates the DNA damage-induced ATM activation through interaction with NBS1

    International Nuclear Information System (INIS)

    Kobayashi, Junya; Tauchi, Hiroshi; Chen, Benjamin; Bruma, Sandeep; Tashiro, Satoshi; Matsuura, Shinya; Tanimoto, Keiji; Chen, David J.; Komatsu, Kenshi

    2009-01-01

    Phosphorylated histone H2AX (γ-H2AX) functions in the recruitment of DNA damage response proteins to DNA double-strand breaks (DSBs) and facilitates DSB repair. ATM also co-localizes with γ-H2AX at DSB sites following its auto-phosphorylation. However, it is unclear whether γ-H2AX has a role in activation of ATM-dependent cell cycle checkpoints. Here, we show that ATM as well as NBS1 is recruited to damaged-chromatin in a γ-H2AX-dependent manner. Foci formation of phosphorylated ATM and ATM-dependent phosphorylation is repressed in H2AX-knockdown cells. Furthermore, anti-γ-H2AX antibody co-immunoprecipitates an ATM-like protein kinase activity in vitro and recombinant H2AX increases in vitro kinase activity of ATM from un-irradiated cells. Moreover, H2AX-deficient cells exhibited a defect in ATM-dependent cell cycle checkpoints. Taken together, γ-H2AX has important role for effective DSB-dependent activation of ATM-related damage responses via NBS1.

  11. Inhibition of the mitotic exit network in response to damaged telomeres.

    Directory of Open Access Journals (Sweden)

    Mauricio Valerio-Santiago

    Full Text Available When chromosomal DNA is damaged, progression through the cell cycle is halted to provide the cells with time to repair the genetic material before it is distributed between the mother and daughter cells. In Saccharomyces cerevisiae, this cell cycle arrest occurs at the G2/M transition. However, it is also necessary to restrain exit from mitosis by maintaining Bfa1-Bub2, the inhibitor of the Mitotic Exit Network (MEN, in an active state. While the role of Bfa1 and Bub2 in the inhibition of mitotic exit when the spindle is not properly aligned and the spindle position checkpoint is activated has been extensively studied, the mechanism by which these proteins prevent MEN function after DNA damage is still unclear. Here, we propose that the inhibition of the MEN is specifically required when telomeres are damaged but it is not necessary to face all types of chromosomal DNA damage, which is in agreement with previous data in mammals suggesting the existence of a putative telomere-specific DNA damage response that inhibits mitotic exit. Furthermore, we demonstrate that the mechanism of MEN inhibition when telomeres are damaged relies on the Rad53-dependent inhibition of Bfa1 phosphorylation by the Polo-like kinase Cdc5, establishing a new key role of this kinase in regulating cell cycle progression.

  12. [Transcranial magnetic stimulation].

    Science.gov (United States)

    Tormos, J M; Catalá, M D; Pascual-Leone, A

    Transcranial magnetic stimulation (TMS) permits stimulation of the cerebral cortex in humans without requiring open access to the brain and is one of the newest tools available in neuroscience. There are two main types of application: single-pulse TMS and repetitive TMS. The magnetic stimulator is composed of a series of capacitors that store the voltage necessary to generate a stimulus of the sufficient intensity of generate an electric field in the stimulation coil. The safety of TMS is supported by the considerable experience derived from studies involving electrical stimulation of the cortex in animals and humans, and also specific studies on the safety of TMS in humans. In this article we review historical and technical aspects of TMS, describe its adverse effects and how to avoid them, summarize the applications of TMS in the investigation of different cerebral functions, and discuss the possibility of using TMS for the treatment of neuropsychiatric disorders.

  13. Structure of a Blinkin-BUBR1 complex reveals an interaction crucial for kinetochore-mitotic checkpoint regulation via an unanticipated binding Site

    DEFF Research Database (Denmark)

    Bolanos-Garcia, Victor M; Lischetti, Tiziana; Matak-Vinković, Dijana

    2011-01-01

    The maintenance of genomic stability relies on the spindle assembly checkpoint (SAC), which ensures accurate chromosome segregation by delaying the onset of anaphase until all chromosomes are properly bioriented and attached to the mitotic spindle. BUB1 and BUBR1 kinases are central for this proc...

  14. Deficiency of the Arabidopsis helicase RTEL1 triggers a SOG1-dependent replication checkpoint in response to DNA cross-links.

    Science.gov (United States)

    Hu, Zhubing; Cools, Toon; Kalhorzadeh, Pooneh; Heyman, Jefri; De Veylder, Lieven

    2015-01-01

    To maintain genome integrity, DNA replication is executed and regulated by a complex molecular network of numerous proteins, including helicases and cell cycle checkpoint regulators. Through a systematic screening for putative replication mutants, we identified an Arabidopsis thaliana homolog of human Regulator of Telomere Length 1 (RTEL1), which functions in DNA replication, DNA repair, and recombination. RTEL1 deficiency retards plant growth, a phenotype including a prolonged S-phase duration and decreased cell proliferation. Genetic analysis revealed that rtel1 mutant plants show activated cell cycle checkpoints, specific sensitivity to DNA cross-linking agents, and increased homologous recombination, but a lack of progressive shortening of telomeres, indicating that RTEL1 functions have only been partially conserved between mammals and plants. Surprisingly, RTEL1 deficiency induces tolerance to the deoxynucleotide-depleting drug hydroxyurea, which could be mimicked by DNA cross-linking agents. This resistance does not rely on the essential replication checkpoint regulator WEE1 but could be blocked by a mutation in the SOG1 transcription factor. Taken together, our data indicate that RTEL1 is required for DNA replication and that its deficiency activates a SOG1-dependent replication checkpoint. © 2015 American Society of Plant Biologists. All rights reserved.

  15. The Level of Europium-154 Contaminating Samarium-153-EDTMP Activates the Radiation Alarm System at the US Homeland Security Checkpoints

    Directory of Open Access Journals (Sweden)

    Mohammed Najeeb Al Hallak

    2009-08-01

    Full Text Available 153Sm-EDTMP is a radiopharmaceutical composed of EDTMP (ethylenediamine-tetramethylenephosphonate and Samarium-153 [1]. 153Sm-EDTMP has an affinity for skeletal tissue and concentrates in areas with increased bone turnover; thus, it is successfully used in relieving pain related to diffuse bone metastases [1]. The manufacturing process of 153Sm-EDTMP leads to contamination with 154Eu (Europium-154 [2]. A previous study only alluded to the retention of 154Eu in the bones after receiving treatment with 153Sm-EDTMP [2]. Activation of the alarm at security checkpoints after 153Sm-EDTMP therapy has not been previously reported. Two out of 15 patients who received 153Sm-EDTMP at Roger Maris Cancer Center (Fargo, N. Dak., USA activated the radiation activity sensors while passing through checkpoints; one at a US airport and the other while crossing theAmerican-Canadian border. We assume that the 154Eu which remained in the patients’ bones activated the sensors. Methods: In order to investigate this hypothesis, we obtained the consent from 3 of our 15 patients who received 153Sm-EDTMP within the previous 4 months to 2 years, including the patient who had activated the radiation alarm at the airport. The patients were scanned with a handheld detector and a gamma camera for energies from 511 keV to 1.3 MeV. Results: All three patients exhibited identical spectral images, and further analysis showed that the observed spectra are the result of 154Eu emissions. Conclusion: Depending on the detection thresholds and windows used by local and federal authorities, the remaining activity of 154Eu retained in patients who received 153Sm-EDTMP could be sufficient enough to increase the count rates above background levels and activate the sensors. At Roger Maris Cancer Center, patients are now informed of the potential consequences of 153Sm-EDTMP therapy prior to initiating treatment. In addition, patients treated with 153Sm-EDTMP at Roger Maris Cancer Center

  16. Alterations of the spindle checkpoint pathway in clinicopathologically aggressive CpG island methylator phenotype clear cell renal cell carcinomas.

    Science.gov (United States)

    Arai, Eri; Gotoh, Masahiro; Tian, Ying; Sakamoto, Hiromi; Ono, Masaya; Matsuda, Akio; Takahashi, Yoriko; Miyata, Sayaka; Totsuka, Hirohiko; Chiku, Suenori; Komiyama, Motokiyo; Fujimoto, Hiroyuki; Matsumoto, Kenji; Yamada, Tesshi; Yoshida, Teruhiko; Kanai, Yae

    2015-12-01

    CpG-island methylator phenotype (CIMP)-positive clear cell renal cell carcinomas (RCCs) are characterized by accumulation of DNA hypermethylation of CpG islands, clinicopathological aggressiveness and poor patient outcome. The aim of this study was to clarify the molecular pathways participating in CIMP-positive renal carcinogenesis. Genome (whole-exome and copy number), transcriptome and proteome (two-dimensional image converted analysis of liquid chromatography-mass spectrometry) analyses were performed using tissue specimens of 87 CIMP-negative and 14 CIMP-positive clear cell RCCs and corresponding specimens of non-cancerous renal cortex. Genes encoding microtubule-associated proteins, such as DNAH2, DNAH5, DNAH10, RP1 and HAUS8, showed a 10% or higher incidence of genetic aberrations (non-synonymous single-nucleotide mutations and insertions/deletions) in CIMP-positive RCCs, whereas CIMP-negative RCCs lacked distinct genetic characteristics. MetaCore pathway analysis of CIMP-positive RCCs revealed that alterations of mRNA or protein expression were significantly accumulated in six pathways, all participating in the spindle checkpoint, including the "The metaphase checkpoint (p = 1.427 × 10(-6))," "Role of Anaphase Promoting Complex in cell cycle regulation (p = 7.444 × 10(-6))" and "Spindle assembly and chromosome separation (p = 9.260 × 10(-6))" pathways. Quantitative RT-PCR analysis revealed that mRNA expression levels for genes included in such pathways, i.e., AURKA, AURKB, BIRC5, BUB1, CDC20, NEK2 and SPC25, were significantly higher in CIMP-positive than in CIMP-negative RCCs. All CIMP-positive RCCs showed overexpression of Aurora kinases, AURKA and AURKB, and this overexpression was mainly attributable to increased copy number. These data suggest that abnormalities of the spindle checkpoint pathway participate in CIMP-positive renal carcinogenesis, and that AURKA and AURKB may be potential therapeutic targets in more aggressive CIMP-positive RCCs.

  17. Music acupuncture stimulation method.

    Science.gov (United States)

    Brătilă, F; Moldovan, C

    2007-01-01

    Harmonic Medicine is the model using the theory that the body rhythms synchronize to an outer rhythm applied for therapeutic purpose, can restores the energy balance in acupuncture channels and organs and the condition of well-being. The purpose of this scientific work was to demonstrate the role played by harmonic sounds in the stimulation of the Lung (LU) Meridian (Shoutaiyin Feijing) and of the Kidney (KI) Meridian (Zushaoyin Shenjing). It was used an original method that included: measurement and electronic sound stimulation of the Meridian Entry Point, measurement of Meridian Exit Point, computer data processing, bio feed-back adjustment of the music stimulation parameters. After data processing, it was found that the sound stimulation of the Lung Meridian Frequency is optimal between 122 Hz and 128 Hz, with an average of 124 Hz (87% of the subjects) and for Kidney Meridian from 118 Hz to 121 Hz, with an average of 120 Hz (67% of the subjects). The acupuncture stimulation was more intense for female subjects (> 7%) than for the male ones. We preliminarily consider that an informational resonance phenomenon can be developed between the acupuncture music stimulation frequency and the cellular dipole frequency, being a really "resonant frequency signature" of an acupoint. The harmonic generation and the electronic excitation or low-excitation status of an acupuncture point may be considered as a resonance mechanism. By this kind of acupunctural stimulation, a symphony may act and play a healer role.

  18. Fission Yeast Apc15 Stabilizes MCC-Cdc20-APC/C Complexes, Ensuring Efficient Cdc20 Ubiquitination and Checkpoint Arrest.

    Science.gov (United States)

    May, Karen M; Paldi, Flora; Hardwick, Kevin G

    2017-04-24

    During mitosis, cells must segregate the replicated copies of their genome to their daughter cells with extremely high fidelity. Segregation errors lead to an abnormal chromosome number (aneuploidy), which typically results in disease or cell death [1]. Chromosome segregation and anaphase onset are initiated through the action of the multi-subunit E3 ubiquitin ligase known as the anaphase-promoting complex or cyclosome (APC/C [2]). The APC/C is inhibited by the spindle checkpoint in the presence of kinetochore attachment defects [3, 4]. Here we demonstrate that two non-essential APC/C subunits (Apc14 and Apc15) regulate association of spindle checkpoint proteins, in the form of the mitotic checkpoint complex (MCC), with the APC/C. apc14Δ mutants display increased MCC association with the APC/C and are unable to silence the checkpoint efficiently. Conversely, apc15Δ mutants display reduced association between the MCC and APC/C, are defective in poly-ubiquitination of Cdc20, and are checkpoint defective. In vitro reconstitution studies have shown that human MCC-APC/C can contain two molecules of Cdc20 [5-7]. Using a yeast strain expressing two Cdc20 genes with different epitope tags, we show by co-immunoprecipitation that this is true in vivo. MCC binding to the second molecule of Cdc20 is mediated via the C-terminal KEN box in Mad3. Somewhat surprisingly, complexes containing both molecules of Cdc20 accumulate in apc15Δ cells, and the implications of this observation are discussed. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  19. Radiation damage to mushrooms

    International Nuclear Information System (INIS)

    Sattler, P.W.

    1986-01-01

    This document contains newspaper cuttings and correspondence with various ministries in Hessen on the subject of radiation damage to mushrooms from the Odenwald area. The reader is given, amongst other things, detailed information on radiation damage to different types of mushroom in 1986. (MG) [de

  20. Animal damage to birch

    Science.gov (United States)

    James S. Jordan; Francis M. Rushmore

    1969-01-01

    A relatively few animal species are responsible for most of the reported damage to the birches. White-tailed deer, yellow-bellied sapsuckers, porcupines, moose, and hares are the major animals involved. We will review reports of damage, discuss the underlying causes, and describe possible methods of control. For example, heavy deer browsing that eliminates birch...

  1. Animal damage management handbook.

    Science.gov (United States)

    Hugh C. Black

    1994-01-01

    This handbook treats animal damage management (ADM) in the West in relation to forest, range, and recreation resources; predator management is not addressed. It provides a comprehensive reference of safe, effective, and practical methods for managing animal damage on National Forest System lands. Supporting information is included in references after each chapter and...

  2. Nuclear damage - civil liability

    International Nuclear Information System (INIS)

    Simoes, A.C.

    1980-01-01

    An analysis is made of the civil liability for nuclear damage since there is a need to adjust the existing rules to the new situations created. The conventions that set up the new disciplining rules not considered in the common law for the liability of nuclear damage are also mentioned. (A.L.) [pt

  3. DNA damage and autophagy

    International Nuclear Information System (INIS)

    Rodriguez-Rocha, Humberto; Garcia-Garcia, Aracely; Panayiotidis, Mihalis I.; Franco, Rodrigo

    2011-01-01

    Both exogenous and endogenous agents are a threat to DNA integrity. Exogenous environmental agents such as ultraviolet (UV) and ionizing radiation, genotoxic chemicals and endogenous byproducts of metabolism including reactive oxygen species can cause alterations in DNA structure (DNA damage). Unrepaired DNA damage has been linked to a variety of human disorders including cancer and neurodegenerative disease. Thus, efficient mechanisms to detect DNA lesions, signal their presence and promote their repair have been evolved in cells. If DNA is effectively repaired, DNA damage response is inactivated and normal cell functioning resumes. In contrast, when DNA lesions cannot be removed, chronic DNA damage triggers specific cell responses such as cell death and senescence. Recently, DNA damage has been shown to induce autophagy, a cellular catabolic process that maintains a balance between synthesis, degradation, and recycling of cellular components. But the exact mechanisms by which DNA damage triggers autophagy are unclear. More importantly, the role of autophagy in the DNA damage response and cellular fate is unknown. In this review we analyze evidence that supports a role for autophagy as an integral part of the DNA damage response.

  4. RPA mediates recombination repair during replication stress and is displaced from DNA by checkpoint signalling in human cells

    DEFF Research Database (Denmark)

    Sleeth, Kate M; Sørensen, Claus Storgaard; Issaeva, Natalia

    2007-01-01

    The replication protein A (RPA) is involved in most, if not all, nuclear metabolism involving single-stranded DNA. Here, we show that RPA is involved in genome maintenance at stalled replication forks by the homologous recombination repair system in humans. Depletion of the RPA protein inhibited...... the formation of RAD51 nuclear foci after hydroxyurea-induced replication stalling leading to persistent unrepaired DNA double-strand breaks (DSBs). We demonstrate a direct role of RPA in homology directed recombination repair. We find that RPA is dispensable for checkpoint kinase 1 (Chk1) activation...... and that RPA directly binds RAD52 upon replication stress, suggesting a direct role in recombination repair. In addition we show that inhibition of Chk1 with UCN-01 decreases dissociation of RPA from the chromatin and inhibits association of RAD51 and RAD52 with DNA. Altogether, our data suggest a direct role...

  5. Human papillomavirus type 16 E7 oncoprotein engages but does not abrogate the mitotic spindle assembly checkpoint

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Yueyang [Division of Infectious Diseases, Brigham and Women' s Hospital and Biological and Biomedical Sciences Program, Harvard Medical School, Boston, MA 02115 (United States); Munger, Karl, E-mail: kmunger@rics.bwh.harvard.edu [Division of Infectious Diseases, Brigham and Women' s Hospital and Biological and Biomedical Sciences Program, Harvard Medical School, Boston, MA 02115 (United States)

    2012-10-10

    The mitotic spindle assembly checkpoint (SAC) ensures faithful chromosome segregation during mitosis by censoring kinetochore-microtubule interactions. It is frequently rendered dysfunctional during carcinogenesis causing chromosome missegregation and genomic instability. There are conflicting reports whether the HPV16 E7 oncoprotein drives chromosomal instability by abolishing the SAC. Here we report that degradation of mitotic cyclins is impaired in cells with HPV16 E7 expression. RNAi-mediated depletion of Mad2 or BubR1 indicated the involvement of the SAC, suggesting that HPV16 E7 expression causes sustained SAC engagement. Mutational analyses revealed that HPV16 E7 sequences that are necessary for retinoblastoma tumor suppressor protein binding as well as sequences previously implicated in binding the nuclear and mitotic apparatus (NuMA) protein and in delocalizing dynein from the mitotic spindle contribute to SAC engagement. Importantly, however, HPV16 E7 does not markedly compromise the SAC response to microtubule poisons.

  6. Requirement of Sequences outside the Conserved Kinase Domain of Fission Yeast Rad3p for Checkpoint Control

    Science.gov (United States)

    Chapman, Carolyn Riley; Evans, Sarah Tyler; Carr, Antony M.; Enoch, Tamar

    1999-01-01

    The fission yeast Rad3p checkpoint protein is a member of the phosphatidylinositol 3-kinase-related family of protein kinases, which includes human ATMp. Mutation of the ATM gene is responsible for the disease ataxia-telangiectasia. The kinase domain of Rad3p has previously been shown to be essential for function. Here, we show that although this domain is necessary, it is not sufficient, because the isolated kinase domain does not have kinase activity in vitro and cannot complement a rad3 deletion strain. Using dominant negative alleles of rad3, we have identified two sites N-terminal to the conserved kinase domain that are essential for Rad3p function. One of these sites is the putative leucine zipper, which is conserved in other phosphatidylinositol 3-kinase-related family members. The other is a novel motif, which may also mediate Rad3p protein–protein interactions. PMID:10512862

  7. Cell-cycle dependent expression of a translocation-mediated fusion oncogene mediates checkpoint adaptation in rhabdomyosarcoma.

    Directory of Open Access Journals (Sweden)

    Ken Kikuchi

    2014-01-01

    Full Text Available Rhabdomyosarcoma is the most commonly occurring soft-tissue sarcoma in childhood. Most rhabdomyosarcoma falls into one of two biologically distinct subgroups represented by alveolar or embryonal histology. The alveolar subtype harbors a translocation-mediated PAX3:FOXO1A fusion gene and has an extremely poor prognosis. However, tumor cells have heterogeneous expression for the fusion gene. Using a conditional genetic mouse model as well as human tumor cell lines, we show that that Pax3:Foxo1a expression is enriched in G2 and triggers a transcriptional program conducive to checkpoint adaptation under stress conditions such as irradiation in vitro and in vivo. Pax3:Foxo1a also tolerizes tumor cells to clinically-established chemotherapy agents and emerging molecularly-targeted agents. Thus, the surprisingly dynamic regulation of the Pax3:Foxo1a locus is a paradigm that has important implications for the way in which oncogenes are modeled in cancer cells.

  8. Leveraging the checkpoint-restart technique for optimizing CPU efficiency of ATLAS production applications on opportunistic platforms

    CERN Document Server

    Cameron, David; The ATLAS collaboration

    2017-01-01

    Data processing applications of the ATLAS experiment, such as event simulation and reconstruction, spend considerable amount of time in the initialization phase. This phase includes loading a large number of shared libraries, reading detector geometry and condition data from external databases, building a transient representation of the detector geometry and initializing various algorithms and services. In some cases the initialization step can take as long as 10-15 minutes. Such slow initialization, being inherently serial, has a significant negative impact on overall CPU efficiency of the production job, especially when the job is executed on opportunistic, often short-lived, resources such as commercial clouds or volunteer computing. In order to improve this situation, we can take advantage of the fact that ATLAS runs large numbers of production jobs with similar configuration parameters (e.g. jobs within the same production task). This allows us to checkpoint one job at the end of its configuration step a...

  9. RB has a critical role in mediating the in vivo checkpoint response, mitigating secondary DNA damage and suppressing liver tumorigenesis initiated by aflatoxin B1

    OpenAIRE

    Reed, CA; Mayhew, CN; McClendon, AK; Yang, X; Witkiewicz, A; Knudsen, ES

    2009-01-01

    Hepatocellular carcinoma (HCC) is a significant worldwide health concern that is associated with discrete etiological events, encompassing viral infection, metabolic stress and genotoxic compounds. In particular, exposure to the genotoxic hepatocarcinogen aflatoxin B1 (AFB1) is a significant factor in the genesis of human liver cancer. Presumably, genetic events associated with HCC could influence the effect of environmental insults, yielding a predilection for tumor development. The retinobl...

  10. Phenotypic characterization of autoreactive B cells--checkpoints of B cell tolerance in patients with systemic lupus erythematosus.

    Directory of Open Access Journals (Sweden)

    Annett M Jacobi

    Full Text Available DNA-reactive B cells play a central role in systemic lupus erythematosus (SLE; DNA antibodies precede clinical disease and in established disease correlate with renal inflammation and contribute to dendritic cell activation and high levels of type 1 interferon. A number of central and peripheral B cell tolerance mechanisms designed to control the survival, differentiation and activation of autoreactive B cells are thought to be disturbed in patients with SLE. The characterization of DNA-reactive B cells has, however, been limited by their low frequency in peripheral blood. Using a tetrameric configuration of a peptide mimetope of DNA bound by pathogenic anti-DNA antibodies, we can identify B cells producing potentially pathogenic DNA-reactive antibodies. We, therefore, characterized the maturation and differentiation states of peptide, (ds double stranded DNA cross-reactive B cells in the peripheral blood of lupus patients and correlated these with clinical disease activity. Flow cytometric analysis demonstrated a significantly higher frequency of tetramer-binding B cells in SLE patients compared to healthy controls. We demonstrated the existence of a novel tolerance checkpoint at the transition of antigen-naïve to antigen-experienced. We further demonstrate that patients with moderately active disease have more autoreactive B cells in both the antigen-naïve and antigen-experienced compartments consistent with greater impairment in B cell tolerance in both early and late checkpoints in these patients than in patients with quiescent disease. This methodology enables us to gain insight into the development and fate of DNA-reactive B cells in individual patients with SLE and paves the way ultimately to permit better and more customized therapies.

  11. Spindle assembly checkpoint protein expression correlates with cellular proliferation and shorter time to recurrence in ovarian cancer.

    LENUS (Irish Health Repository)

    McGrogan, Barbara

    2014-07-01

    Ovarian carcinoma (OC) is the most lethal of the gynecological malignancies, often presenting at an advanced stage. Treatment is hampered by high levels of drug resistance. The taxanes are microtubule stabilizing agents, used as first-line agents in the treatment of OC that exert their apoptotic effects through the spindle assembly checkpoint. BUB1-related protein kinase (BUBR1) and mitotic arrest deficient 2 (MAD2), essential spindle assembly checkpoint components, play a key role in response to taxanes. BUBR1, MAD2, and Ki-67 were assessed on an OC tissue microarray platform representing 72 OC tumors of varying histologic subtypes. Sixty-one of these patients received paclitaxel and platinum agents combined; 11 received platinum alone. Overall survival was available for all 72 patients, whereas recurrence-free survival (RFS) was available for 66 patients. Increased BUBR1 expression was seen in serous carcinomas, compared with other histologies (P = .03). Increased BUBR1 was significantly associated with tumors of advanced stage (P = .05). Increased MAD2 and BUBR1 expression also correlated with increased cellular proliferation (P < .0002 and P = .02, respectively). Reduced MAD2 nuclear intensity was associated with a shorter RFS (P = .03), in ovarian tumors of differing histologic subtype (n = 66). In this subgroup, for those women who received paclitaxel and platinum agents combined (n = 57), reduced MAD2 intensity also identified women with a shorter RFS (P < .007). For the entire cohort of patients, irrespective of histologic subtype or treatment, MAD2 nuclear intensity retained independent significance in a multivariate model, with tumors showing reduced nuclear MAD2 intensity identifying patients with a poorer RFS (P = .05).

  12. Metabolite Damage and Metabolite Damage Control in Plants

    Energy Technology Data Exchange (ETDEWEB)

    Hanson, Andrew D. [Horticultural Sciences Department and; Henry, Christopher S. [Mathematics and Computer Science Division, Argonne National Laboratory, Argonne, Illinois 60439, email:; Computation Institute, University of Chicago, Chicago, Illinois 60637; Fiehn, Oliver [Genome Center, University of California, Davis, California 95616, email:; de Crécy-Lagard, Valérie [Microbiology and Cell Science Department, University of Florida, Gainesville, Florida 32611, email: ,

    2016-04-29

    It is increasingly clear that (a) many metabolites undergo spontaneous or enzyme-catalyzed side reactions in vivo, (b) the damaged metabolites formed by these reactions can be harmful, and (c) organisms have biochemical systems that limit the buildup of damaged metabolites. These damage-control systems either return a damaged molecule to its pristine state (metabolite repair) or convert harmful molecules to harmless ones (damage preemption). Because all organisms share a core set of metabolites that suffer the same chemical and enzymatic damage reactions, certain damage-control systems are widely conserved across the kingdoms of life. Relatively few damage reactions and damage-control systems are well known. Uncovering new damage reactions and identifying the corresponding damaged metabolites, damage-control genes, and enzymes demands a coordinated mix of chemistry, metabolomics, cheminformatics, biochemistry, and comparative genomics. This review illustrates the above points using examples from plants, which are at least as prone to metabolite damage as other organisms.

  13. Optical stimulation of the facial nerve: a surgical tool?

    Science.gov (United States)

    Richter, Claus-Peter; Teudt, Ingo Ulrik; Nevel, Adam E.; Izzo, Agnella D.; Walsh, Joseph T., Jr.

    2008-02-01

    One sequela of skull base surgery is the iatrogenic damage to cranial nerves. Devices that stimulate nerves with electric current can assist in the nerve identification. Contemporary devices have two main limitations: (1) the physical contact of the stimulating electrode and (2) the spread of the current through the tissue. In contrast to electrical stimulation, pulsed infrared optical radiation can be used to safely and selectively stimulate neural tissue. Stimulation and screening of the nerve is possible without making physical contact. The gerbil facial nerve was irradiated with 250-μs-long pulses of 2.12 μm radiation delivered via a 600-μm-diameter optical fiber at a repetition rate of 2 Hz. Muscle action potentials were recorded with intradermal electrodes. Nerve samples were examined for possible tissue damage. Eight facial nerves were stimulated with radiant exposures between 0.71-1.77 J/cm2, resulting in compound muscle action potentials (CmAPs) that were simultaneously measured at the m. orbicularis oculi, m. levator nasolabialis, and m. orbicularis oris. Resulting CmAP amplitudes were 0.3-0.4 mV, 0.15-1.4 mV and 0.3-2.3 mV, respectively, depending on the radial location of the optical fiber and the radiant exposure. Individual nerve branches were also stimulated, resulting in CmAP amplitudes between 0.2 and 1.6 mV. Histology revealed tissue damage at radiant exposures of 2.2 J/cm2, but no apparent damage at radiant exposures of 2.0 J/cm2.

  14. The cell-cycle checkpoint kinase Chk1 is required for mammalian homologous recombination repair

    DEFF Research Database (Denmark)

    Sørensen, Claus Storgaard; Hansen, Lasse Tengbjerg; Dziegielewski, Jaroslaw

    2005-01-01

    repair (HRR) system. Abrogation of Chk1 function with small interfering RNA or chemical antagonists inhibits HRR, leading to persistent unrepaired DNA double-strand breaks (DSBs) and cell death after replication inhibition with hydroxyurea or DNA-damage caused by camptothecin. After hydroxyurea treatment......-depleted cells failed to form RAD51 nuclear foci after exposure to hydroxyurea, and cells expressing a phosphorylation-deficient mutant RAD51(T309A) were hypersensitive to hydroxyurea. These results highlight a crucial role for the Chk1 signalling pathway in protecting cells against lethal DNA lesions...

  15. The role of p53 in the response to mitotic spindle damage

    International Nuclear Information System (INIS)

    Meek, D.W.

    2000-01-01

    The p53 tumour suppressor protein has defined roles in G1/S and G2/M cell cycle checkpoint in response to a range of cellular stresses including DNA damage, dominant oncogene expression, hypoxia, metabolic changes and viral infection. In addition to these responses, p53 can also be activated when damage occurs to the mitotic spindle. Initially, spindle damage activates a p53-independent checkpoint which functions at the metaphase-anaphase transition and prevents cells from progressing through mitosis until the completion of spindle formation. Cells eventually escape from this block (a process termed 'mitotic slippage'), and an aberrant mitosis ensues in which sister chromatids fail to segregate properly. After a delay period, p53 responds to this mitotic failure by instituting a G1-like growth arrest, with an intact nucleus containing 4N DNA, but without the cells undergoing division. Cells lacking wild-type p53 are still able to arrest transiently at mitosis, and also fail to undergo division, underscoring that the delay in mitosis is p53-independent. However, these cells are not prevented from re-entering the cell cycle and can reduplicate their DNA unchecked, leading to polyploidy. Additionally, p53-null cells which experience spindle failure often show the appearance of micronuclei arising from poorly segregated chromosomes which have de-condensed and been enclosed in a nuclear envelope. The ability of p53 to prevent their formation suggests an additional G2 involvement which prevents nuclear breakdown prior to mitosis. The molecular mechanism by which p53 is able to sense mitotic failure is still unknown, but may be linked to the ability of p53 to regulate duplication of the centrosome, the organelle which nucleates spindle formation. (authors)

  16. Apoptosis-like yeast cell death in response to DNA damage and replication defects

    Energy Technology Data Exchange (ETDEWEB)

    Burhans, William C.; Weinberger, Martin; Marchetti, Maria A.; Ramachandran, Lakshmi; D' Urso, Gennaro; Huberman, Joel A

    2003-11-27

    In budding (Saccharomyces cerevisiae) and fission (Schizosaccharomyces pombe) yeast and other unicellular organisms, DNA damage and other stimuli can induce cell death resembling apoptosis in metazoans, including the activation of a recently discovered caspase-like molecule in budding yeast. Induction of apoptotic-like cell death in yeasts requires homologues of cell cycle checkpoint proteins that are often required for apoptosis in metazoan cells. Here, we summarize these findings and our unpublished results which show that an important component of metazoan apoptosis recently detected in budding yeast - reactive oxygen species (ROS) - can also be detected in fission yeast undergoing an apoptotic-like cell death. ROS were detected in fission and budding yeast cells bearing conditional mutations in genes encoding DNA replication initiation proteins and in fission yeast cells with mutations that deregulate cyclin-dependent kinases (CDKs). These mutations may cause DNA damage by permitting entry of cells into S phase with a reduced number of replication forks and/or passage through mitosis with incompletely replicated chromosomes. This may be relevant to the frequent requirement for elevated CDK activity in mammalian apoptosis, and to the recent discovery that the initiation protein Cdc6 is destroyed during apoptosis in mammals and in budding yeast cells exposed to lethal levels of DNA damage. Our data indicate that connections between apoptosis-like cell death and DNA replication or CDK activity are complex. Some apoptosis-like pathways require checkpoint proteins, others are inhibited by them, and others are independent of them. This complexity resembles that of apoptotic pathways in mammalian cells, which are frequently deregulated in cancer. The greater genetic tractability of yeasts should help to delineate these complex pathways and their relationships to cancer and to the effects of apoptosis-inducing drugs that inhibit DNA replication.

  17. Apoptosis-like yeast cell death in response to DNA damage and replication defects

    International Nuclear Information System (INIS)

    Burhans, William C.; Weinberger, Martin; Marchetti, Maria A.; Ramachandran, Lakshmi; D'Urso, Gennaro; Huberman, Joel A.

    2003-01-01

    In budding (Saccharomyces cerevisiae) and fission (Schizosaccharomyces pombe) yeast and other unicellular organisms, DNA damage and other stimuli can induce cell death resembling apoptosis in metazoans, including the activation of a recently discovered caspase-like molecule in budding yeast. Induction of apoptotic-like cell death in yeasts requires homologues of cell cycle checkpoint proteins that are often required for apoptosis in metazoan cells. Here, we summarize these findings and our unpublished results which show that an important component of metazoan apoptosis recently detected in budding yeast - reactive oxygen species (ROS) - can also be detected in fission yeast undergoing an apoptotic-like cell death. ROS were detected in fission and budding yeast cells bearing conditional mutations in genes encoding DNA replication initiation proteins and in fission yeast cells with mutations that deregulate cyclin-dependent kinases (CDKs). These mutations may cause DNA damage by permitting entry of cells into S phase with a reduced number of replication forks and/or passage through mitosis with incompletely replicated chromosomes. This may be relevant to the frequent requirement for elevated CDK activity in mammalian apoptosis, and to the recent discovery that the initiation protein Cdc6 is destroyed during apoptosis in mammals and in budding yeast cells exposed to lethal levels of DNA damage. Our data indicate that connections between apoptosis-like cell death and DNA replication or CDK activity are complex. Some apoptosis-like pathways require checkpoint proteins, others are inhibited by them, and others are independent of them. This complexity resembles that of apoptotic pathways in mammalian cells, which are frequently deregulated in cancer. The greater genetic tractability of yeasts should help to delineate these complex pathways and their relationships to cancer and to the effects of apoptosis-inducing drugs that inhibit DNA replication

  18. Cellular Inhibition of Checkpoint Kinase 2 (Chk2) and Potentiation of Camptothecins and Radiation by the Novel Chk2 Inhibitor PV1019 [7-Nitro-1H-indole-2-carboxylic acid {4-[1-(guanidinohydrazone)-ethyl]-phenyl}-amide

    Energy Technology Data Exchange (ETDEWEB)

    Jobson, Andrew G.; Lountos, George T.; Lorenzi, Philip L.; Llamas, Jenny; Connelly, John; Cerna, David; Tropea, Joseph E.; Onda, Akikazu; Zoppoli, Gabriele; Kondapaka, Sudhir; Zhang, Guangtao; Caplen, Natasha J.; Cardellina, II, John H.; Yoo, Stephen S.; Monks, Anne; Self, Christopher; Waugh, David S.; Shoemaker, Robert H.; Pommier, Yves; (NIH)

    2010-04-05

    Chk2 is a checkpoint kinase involved in the ataxia telangiectasia mutated pathway, which is activated by genomic instability and DNA damage, leading to either cell death (apoptosis) or cell cycle arrest. Chk2 provides an unexplored therapeutic target against cancer cells. We recently reported 4,4'-diacetyldiphenylurea-bis(guanylhydrazone) (NSC 109555) as a novel chemotype Chk2 inhibitor. We have now synthesized a derivative of NSC 109555, PV1019 (NSC 744039) [7-nitro-1H-indole-2-carboxylic acid {l_brace}4-[1-(guanidinohydrazone)-ethyl]-phenyl{r_brace}-amide], which is a selective submicromolar inhibitor of Chk2 in vitro. The cocrystal structure of PV1019 bound in the ATP binding pocket of Chk2 confirmed enzymatic/biochemical observations that PV1019 acts as a competitive inhibitor of Chk2 with respect to ATP. PV1019 was found to inhibit Chk2 in cells. It inhibits Chk2 autophosphorylation (which represents the cellular kinase activation of Chk2), Cdc25C phosphorylation, and HDMX degradation in response to DNA damage. PV1019 also protects normal mouse thymocytes against ionizing radiation-induced apoptosis, and it shows synergistic antiproliferative activity with topotecan, camptothecin, and radiation in human tumor cell lines. We also show that PV1019 and Chk2 small interfering RNAs can exert antiproliferative activity themselves in the cancer cells with high Chk2 expression in the NCI-60 screen. These data indicate that PV1019 is a potent and selective inhibitor of Chk2 with chemotherapeutic and radiosensitization potential.

  19. Replication-mediated disassociation of replication protein A-XPA complex upon DNA damage: implications for RPA handing off.

    Science.gov (United States)

    Jiang, Gaofeng; Zou, Yue; Wu, Xiaoming

    2012-08-01

    RPA (replication protein A), the eukaryotic ssDNA (single-stranded DNA)-binding protein, participates in most cellular processes in response to genotoxic insults, such as NER (nucleotide excision repair), DNA, DSB (double-strand break) repair and activation of cell cycle checkpoint signalling. RPA interacts with XPA (xeroderma pigmentosum A) and functions in early stage of NER. We have shown that in cells the RPA-XPA complex disassociated upon exposure of cells to high dose of UV irradiation. The dissociation required replication stress and was partially attributed to tRPA hyperphosphorylation. Treatment of cells with CPT (camptothecin) and HU (hydroxyurea), which cause DSB DNA damage and replication fork collapse respectively and also leads to the disruption of RPA-XPA complex. Purified RPA and XPA were unable to form complex in vitro in the presence of ssDNA. We propose that the competition-based RPA switch among different DNA metabolic pathways regulates the dissociation of RPA with XPA in cells after DNA damage. The biological significances of RPA-XPA complex disruption in relation with checkpoint activation, DSB repair and RPA hyperphosphorylation are discussed.

  20. Replication-mediated disassociation of replication protein A–XPA complex upon DNA damage: implications for RPA handing off

    Science.gov (United States)

    Jiang, Gaofeng; Zou, Yue; Wu, Xiaoming

    2013-01-01

    RPA (replication protein A), the eukaryotic ssDNA (single-stranded DNA)-binding protein, participates in most cellular processes in response to genotoxic insults, such as NER (nucleotide excision repair), DNA, DSB (double-strand break) repair and activation of cell cycle checkpoint signalling. RPA interacts with XPA (xeroderma pigmentosum A) and functions in early stage of NER. We have shown that in cells the RPA–XPA complex disassociated upon exposure of cells to high dose of UV irradiation. The dissociation required replication stress and was partially attributed to tRPA hyperphosphorylation. Treatment of cells with CPT (camptothecin) and HU (hydroxyurea), which cause DSB DNA damage and replication fork collapse respectively and also leads to the disruption of RPA–XPA complex. Purified RPA and XPA were unable to form complex in vitro in the presence of ssDNA. We propose that the competition-based RPA switch among different DNA metabolic pathways regulates the dissociation of RPA with XPA in cells after DNA damage. The biological significances of RPA–XPA complex disruption in relation with checkpoint activation, DSB repair and RPA hyperphosphorylation are discussed. PMID:22578086

  1. Electrical and optical co-stimulation in the deaf white cat

    Science.gov (United States)

    Cao, Zhiping; Xu, Yingyue; Tan, Xiaodong; Suematsu, Naofumi; Robinson, Alan; Richter, Claus-Peter

    2018-02-01

    Spatial selectivity of neural stimulation with photons, such as infrared neural stimulation (INS) is higher than the selectivity obtained with electrical stimulation. To obtain more independent channels for stimulation in neural prostheses, INS may be implemented to better restore the fidelity of the damaged neural system. However, irradiation with infrared light also bares the risk of heat accumulation in the target tissue with subsequent neural damage. Lowering the threshold for stimulation could reduce the amount of heat delivered to the tissue and the risk for subsequent tissue damage. It has been shown in the rat sciatic nerve that simultaneous irradiation with infrared light and the delivery of biphasic sub-threshold electrical pulses can reduce the threshold for INS [1]. In this study, deaf white cats have been used to test whether opto-electrical co-stimulation can reduce the stimulation threshold for INS in the auditory system too. The cochleae of the deaf white cats have largely reduced spiral ganglion neuron counts and significant degeneration of the organ of Corti and do not respond to acoustic stimuli. Combined electrical and optical stimulation was used to demonstrate that simultaneous stimulation with infrared light and biphasic electrical pulses can reduce the threshold for stimulation.

  2. LSD and Genetic Damage

    Science.gov (United States)

    Dishotsky, Norman I.; And Others

    1971-01-01

    Reviews studies of the effects of lysergic acid diethylamide (LSD) on man and other organisms. Concludes that pure LSD injected in moderate doses does not cause chromosome or detectable genetic damage and is not a teratogen or carcinogen. (JM)

  3. Diabetes and nerve damage

    Science.gov (United States)

    Diabetic neuropathy; Diabetes - neuropathy; Diabetes - peripheral neuropathy ... In people with diabetes, the body's nerves can be damaged by decreased blood flow and a high blood sugar level. This condition is ...

  4. Stimulated Brillouin processes in crystals and glasses

    International Nuclear Information System (INIS)

    Faris, G.W.; Hickman, A.P.

    1992-02-01

    The basic physics and material properties needed to describe and predict the Brillouin gain for a variety of materials have been investigated. Lawrence Livermore National Laboratory (LLNL) has identified transverse stimulated Brillouin scattering (SBS) as an important limiting mechanism in high power laser fusion systems. At sufficiently high laser intensities, SBS drives acoustic vibrations that can damage optical components. SRI has performed measurements and developed the corresponding theory for stimulated Brillouin gain spectroscopy in anisotropic crystals. Absolute Brillouin steady-state gain coefficients, linewidths, and frequency shifts have been determined at 532 nm for a number of optical materials of interest to LLNL. This knowledge can be used to select optical materials and devise suppression schemes that will allow much higher laser fluences to be used in laser fusion

  5. New York Canyon Stimulation

    Energy Technology Data Exchange (ETDEWEB)

    Raemy, Bernard

    2012-06-21

    The New York Canyon Stimulation Project was to demonstrate the commercial application of Enhanced Geothermal System techniques in Buena Vista Valley area of Pershing County, Nevada. From October 2009 to early 2012, TGP Development Company aggressively implemented Phase I of Pre-Stimulation and Site/Wellbore readiness. This included: geological studies; water studies and analyses and procurement of initial permits for drilling. Oversubscription of water rights and lack of water needed for implementation of EGS were identified and remained primary obstacles. Despite extended efforts to find alternative solutions, the water supply circumstances could not be overcome and led TGP to determine a "No Go" decision and initiate project termination in April 2012.

  6. A case of pembrolizumab-induced type-1 diabetes mellitus and discussion of immune checkpoint inhibitor-induced type 1 diabetes.

    Science.gov (United States)

    Chae, Young Kwang; Chiec, Lauren; Mohindra, Nisha; Gentzler, Ryan; Patel, Jyoti; Giles, Francis

    2017-01-01

    Immune checkpoint inhibitors such as pembrolizumab, ipilimumab, and nivolumab, now FDA-approved for use in treating several types of cancer, have been associated with immune-related adverse effects. Specifically, the antibodies targeting the programmed-cell death-1 immune checkpoint, pembrolizumab and nivolumab, have been rarely reported to induce the development of type 1 diabetes mellitus. Here we describe a case of a patient who developed antibody-positive type 1 diabetes mellitus following treatment with pembrolizumab in combination with systemic chemotherapy for metastatic adenocarcinoma of the lung. We will also provide a brief literature review of other rarely reported cases of type 1 diabetes presenting after treatment with pembrolizumab and nivolumab, as well as discussion regarding potential mechanisms of this adverse effect and its importance as these drugs continue to become even more widespread.

  7. Incorporation of Immune Checkpoint Blockade into Chimeric Antigen Receptor T Cells (CAR-Ts): Combination or Built-In CAR-T.

    Science.gov (United States)

    Yoon, Dok Hyun; Osborn, Mark J; Tolar, Jakub; Kim, Chong Jai

    2018-01-24

    Chimeric antigen receptor (CAR) T cell therapy represents the first U.S. Food and Drug Administration approved gene therapy and these engineered cells function with unprecedented efficacy in the treatment of refractory CD19 positive hematologic malignancies. CAR translation to solid tumors is also being actively investigated; however, efficacy to date has been variable due to tumor-evolved mechanisms that inhibit local immune cell activity. To bolster the potency of CAR-T cells, modulation of the immunosuppressive tumor microenvironment with immune-checkpoint blockade is a promising strategy. The impact of this approach on hematological malignancies is in its infancy, and in this review we discuss CAR-T cells and their synergy with immune-checkpoint blockade.

  8. Checkpoint-based forward recovery using lookahead execution and rollback validation in parallel and distributed systems. Ph.D. Thesis, 1992

    Science.gov (United States)

    Long, Junsheng

    1994-01-01

    This thesis studies a forward recovery strategy using checkpointing and optimistic execution in parallel and distributed systems. The approach uses replicated tasks executing on different processors for forwared recovery and checkpoint comparison for error detection. To reduce overall redundancy, this approach employs a lower static redundancy in the common error-free situation to detect error than the standard N Module Redundancy scheme (NMR) does to mask off errors. For the rare occurrence of an error, this approach uses some extra redundancy for recovery. To reduce the run-time recovery overhead, look-ahead processes are used to advance computation speculatively and a rollback process is used to produce a diagnosis for correct look-ahead processes without rollback of the whole system. Both analytical and experimental evaluation have shown that this strategy can provide a nearly error-free execution time even under faults with a lower average redundancy than NMR.

  9. Incorporation of Immune Checkpoint Blockade into Chimeric Antigen Receptor T Cells (CAR-Ts: Combination or Built-In CAR-T

    Directory of Open Access Journals (Sweden)

    Dok Hyun Yoon

    2018-01-01

    Full Text Available Chimeric antigen receptor (CAR T cell therapy represents the first U.S. Food and Drug Administration approved gene therapy and these engineered cells function with unprecedented efficacy in the treatment of refractory CD19 positive hematologic malignancies. CAR translation to solid tumors is also being actively investigated; however, efficacy to date has been variable due to tumor-evolved mechanisms that inhibit local immune cell activity. To bolster the potency of CAR-T cells, modulation of the immunosuppressive tumor microenvironment with immune-checkpoint blockade is a promising strategy. The impact of this approach on hematological malignancies is in its infancy, and in this review we discuss CAR-T cells and their synergy with immune-checkpoint blockade.

  10. The Possible Crosstalk of MOB2 With NDR1/2 Kinases in Cell Cycle and DNA Damage Signaling.

    Science.gov (United States)

    Gundogdu, Ramazan; Hergovich, Alexander

    2016-09-06

    This article is the authors' opinion of the roles of the signal transducer Mps one binder 2 (MOB2) in the control of cell cycle progression and the DNA Damage Response (DDR). We recently found that endogenous MOB2 is required to prevent the accumulation of endogenous DNA damage in order to prevent the undesired, and possibly detrimental, activation of cell cycle checkpoints. In this regard, it is noteworthy that MOB2 has been linked biochemically to the regulation of the NDR1/2 (aka STK38/STK38L) protein kinases, which themselves have functions at different steps of the cell cycle. Therefore, we are speculating in this article about the possible connections of MOB2 with NDR1/2 kinases in cell cycle and DDR Signaling.

  11. IDEA: Stimulating Oral Production.

    Science.gov (United States)

    Easley, Jacob J.

    1995-01-01

    Presents daily activities that facilitate complete sentence response, promote oral production, and aid the learning of vocabulary in foreign-language classes. Because speech is the primary form of communication in the foreign-language classroom, it is important to stimulate students to converse as soon as possible. (Author/CK)

  12. stimulated BV2 Microglial

    African Journals Online (AJOL)

    2012-03-26

    Mar 26, 2012 ... 2), in LPS-stimulated BV2 microglial cells. The level of NO production was analyzed using Griess reaction. The release of PGE2 was determined using sandwich enzyme-linked immunosorbent assay. The DNA-binding activity of nuclear factor-κB (NF-κB) was measured by electrophoretic mobility shift assay ...

  13. Brain stimulation in migraine.

    Science.gov (United States)

    Brighina, Filippo; Cosentino, Giuseppe; Fierro, Brigida

    2013-01-01

    Migraine is a very prevalent disease with great individual disability and socioeconomic burden. Despite intensive research effort in recent years, the etiopathogenesis of the disease remains to be elucidated. Recently, much importance has been given to mechanisms underlying the cortical excitability that has been suggested to be dysfunctional in migraine. In recent years, noninvasive brain stimulation techniques based on magnetic fields (transcranial magnetic stimulation, TMS) and on direct electrical currents (transcranial direct current stimulation, tDCS) have been shown to be safe and effective tools to explore the issue of cortical excitability, activation, and plasticity in migraine. Moreover, TMS, repetitive TMS (rTMS), and tDCS, thanks to their ability to interfere with and/or modulate cortical activity inducing plastic, persistent effects, have been also explored as potential therapeutic approaches, opening an interesting perspective for noninvasive neurostimulation for both symptomatic and preventive treatment of migraine and other types of headache. In this chapter we critically review evidence regarding the role of noninvasive brain stimulation in the pathophysiology and treatment of migraine, delineating the advantages and limits of these techniques together with potential development and future application. © 2013 Elsevier B.V. All rights reserved.

  14. A Phase 1 Trial of an Immune Checkpoint Inhibitor plus Stereotactic Ablative Radiotherapy in Patients with Inoperable Stage I Non-Small Cell Lung Cancer

    Science.gov (United States)

    2017-10-01

    with Inoperable Stage I Non-Small Cell Lung Cancer PRINCIPAL INVESTIGATOR: Karen Kelly, MD CONTRACTING ORGANIZATION: University of California...Inhibitor plus Stereotactic Ablative Radiotherapy in Patients with Inoperable Stage I Non-Small Cell Lung Cancer 5b. GRANT NUMBER W81XWH-15-2-0063...immune checkpoint inhibitor MPDL3280A (atezolizumab) in early stage inoperable non-small cell lung cancer . The trial is comprised of a traditional 3 + 3

  15. A Novel Antagonist of the Immune Checkpoint Protein Adenosine A2a Receptor Restores Tumor-Infiltrating Lymphocyte Activity in the Context of the Tumor Microenvironment

    Directory of Open Access Journals (Sweden)

    Melanie Mediavilla-Varela

    2017-07-01

    Full Text Available BACKGROUND: Therapeutic strategies targeting immune checkpoint proteins have led to significant responses in patients with various tumor types. The success of these studies has led to the development of various antibodies/inhibitors for the different checkpoint proteins involved in immune evasion of the tumor. Adenosine present in high concentrations in the tumor microenvironment activates the immune checkpoint adenosine A2a receptor (A2aR, leading to the suppression of antitumor responses. Inhibition of this checkpoint has the potential to enhance antitumor T-cell responsiveness. METHODS: We developed a novel A2aR antagonist (PBF-509 and tested its antitumor response in vitro, in a mouse model, and in non-small cell lung cancer patient samples. RESULTS: Our studies showed that PBF-509 is highly specific to the A2aR as well as inhibitory of A2aR function in an in vitro model. In a mouse model, we found that lung metastasis was decreased after treatment with PBF-509 compared with its control. Furthermore, freshly resected tumor-infiltrating lymphocytes from lung cancer patients showed increased A2aR expression in CD4+ cells and variable expression in CD8+ cells. Ex vivo studies showed an increased responsiveness of human tumor-infiltrating lymphocytes when PBF-509 was combined with anti-PD-1 or anti-PD-L1. CONCLUSIONS: Our studies demonstrate that inhibition of the A2aR using the novel inhibitor PBF-509 could lead to novel immunotherapeutic strategies in non-small cell lung cancer.

  16. Solar ultraviolet radiation-induced DNA damage in aquatic organisms: potential environmental impact

    International Nuclear Information System (INIS)

    Haeder, Donat-P.; Sinha, Rajeshwar P.

    2005-01-01

    Continuing depletion of stratospheric ozone and subsequent increases in deleterious ultraviolet (UV) radiation at the Earth's surface have fueled the interest in its ecological consequences for aquatic ecosystems. The DNA is certainly one of the key targets for UV-induced damage in a variety of aquatic organisms. UV radiation induces two of the most abundant mutagenic and cytotoxic DNA lesions, cyclobutane pyrimidine dimers (CPDs) and pyrimidine pyrimidone photoproducts (6-4PPs) and their Dewar valence isomers. However, aquatic organisms have developed a number of repair and tolerance mechanisms to counteract the damaging effects of UV on DNA. Photoreactivation with the help of the enzyme photolyase is one of the most important and frequently occurring repair mechanisms in a variety of organisms. Excision repair, which can be distinguished into base excision repair (BER) and nucleotide excision repair (NER), also play an important role in DNA repair in several organisms with the help of a number of glycosylases and polymerases, respectively. In addition, mechanisms such as mutagenic repair or dimer bypass, recombinational repair, cell-cycle checkpoints, apoptosis and certain alternative repair pathways are also operative in various organisms. This review deals with the UV-induced DNA damage and repair in a number of aquatic organisms as well as methods of detecting DNA damage

  17. Controlling the response to DNA damage by the APC/C-Cdh1.

    Science.gov (United States)

    de Boer, H Rudolf; Guerrero Llobet, S; van Vugt, Marcel A T M

    2016-03-01

    Proper cell cycle progression is safeguarded by the oscillating activities of cyclin/cyclin-dependent kinase complexes. An important player in the regulation of mitotic cyclins is the anaphase-promoting complex/cyclosome (APC/C), a multi-subunit E3 ubiquitin ligase. Prior to entry into mitosis, the APC/C remains inactive, which allows the accumulation of mitotic regulators. APC/C activation requires binding to either the Cdc20 or Cdh1 adaptor protein, which sequentially bind the APC/C and facilitate targeting of multiple mitotic regulators for proteasomal destruction, including Securin and Cyclin B, to ensure proper chromosome segregation and mitotic exit. Emerging data have indicated that the APC/C, particularly in association with Cdh1, also functions prior to mitotic entry. Specifically, the APC/C-Cdh1 is activated in response to DNA damage in G2 phase cells. These observations are in line with in vitro and in vivo genetic studies, in which cells lacking Cdh1 expression display various defects, including impaired DNA repair and aberrant cell cycle checkpoints. In this review, we summarize the current literature on APC/C regulation in response to DNA damage, the functions of APC/C-Cdh1 activation upon DNA damage, and speculate how APC/C-Cdh1 can control cell fate in the context of persistent DNA damage.

  18. Aven-mediated checkpoint kinase control regulates proliferation and resistance to chemotherapy in conventional osteosarcoma.

    Science.gov (United States)

    Baranski, Zuzanna; Booij, Tijmen H; Cleton-Jansen, Anne-Marie; Price, Leo S; van de Water, Bob; Bovée, Judith V M G; Hogendoorn, Pancras C W; Danen, Erik H J

    2015-07-01

    Conventional high-grade osteosarcoma is the most common primary bone sarcoma, with relatively high incidence in young people. In this study we found that expression of Aven correlates inversely with metastasis-free survival in osteosarcoma patients and is increased in metastases compared to primary tumours. Aven is an adaptor protein that has been implicated in anti-apoptotic signalling and serves as an oncoprotein in acute lymphoblastic leukaemia. In osteosarcoma cells, silencing Aven triggered G2 cell-cycle arrest; Chk1 protein levels were attenuated and ATR-Chk1 DNA damage response signalling in response to chemotherapy was abolished in Aven-depleted osteosarcoma cells, while ATM, Chk2 and p53 activation remained intact. Osteosarcoma is notoriously difficult to treat with standard chemotherapy, and we examined whether pharmacological inhibition of the Aven-controlled ATR-Chk1 response could sensitize osteosarcoma cells to genotoxic compounds. Indeed, pharmacological inhibitors targeting Chk1/Chk2 or those selective for Chk1 synergized with standard chemotherapy in 2D cultures. Likewise, in 3D extracellular matrix-embedded cultures, Chk1 inhibition led to effective sensitization to chemotherapy. Together, these findings implicate Aven in ATR-Chk1 signalling and point towards Chk1 inhibition as a strategy to sensitize human osteosarcomas to chemotherapy. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  19. Oncogenic Herpesvirus Utilizes Stress-Induced Cell Cycle Checkpoints for Efficient Lytic Replication.

    Directory of Open Access Journals (Sweden)

    Giuseppe Balistreri

    2016-02-01

    Full Text Available Kaposi's sarcoma herpesvirus (KSHV causes Kaposi's sarcoma and certain lymphoproliferative malignancies. Latent infection is established in the majority of tumor cells, whereas lytic replication is reactivated in a small fraction of cells, which is important for both virus spread and disease progression. A siRNA screen for novel regulators of KSHV reactivation identified the E3 ubiquitin ligase MDM2 as a negative regulator of viral reactivation. Depletion of MDM2, a repressor of p53, favored efficient activation of the viral lytic transcription program and viral reactivation. During lytic replication cells activated a p53 response, accumulated DNA damage and arrested at G2-phase. Depletion of p21, a p53 target gene, restored cell cycle progression and thereby impaired the virus reactivation cascade delaying the onset of virus replication induced cytopathic effect. Herpesviruses are known to reactivate in response to different kinds of stress, and our study now highlights the molecular events in the stressed host cell that KSHV has evolved to utilize to ensure efficient viral lytic replication.

  20. Attenuation of G2 cell cycle checkpoint control in human tumor cells is associated with increased frequencies of unrejoined chromosome breaks but not increased cytotoxicity following radiation exposure

    International Nuclear Information System (INIS)

    Schwartz, J.L.; Cowan, J.; Grdina, D.J.

    1997-01-01

    The contribution of G 2 cell cycle checkpoint control to ionizing radiation responses was examined in ten human tumor cell lines. Most of the delay in cell cycle progression seen in the first cell cycle following radiation exposure was due to blocks in G 2 and there were large cell line-to-cell line variations in the length of the G 2 block. Longer delays were seen in cell lines that had mutations in p53. There was a highly significant inverse correlation between the length of G 2 delay and the frequency of unrejoined chromosome breaks seen as chromosome terminal deletions in mitosis, and observation that supports the hypothesis that the signal for G 2 delay in mammalian cells is an unrejoined chromosome break. There were also an inverse correlation between the length of G 2 delay and the level of chromosome aneuploidy in each cell line, suggesting that the G 2 and mitotic spindel checkpoints may be linked to each other. Attenuation in G 2 checkpoint control was not associated with alterations in either the frequency of induced chromosome rearrangements or cell survival following radiation exposure suggesting that chromosome rearrangements, the major radiation-induced lethal lesion in tumor cells, form before cells enters G 2 . Thus, agents that act solely to override G 2 arrest should produce little radiosensitization in human tumor cells

  1. Coal transportation road damage

    International Nuclear Information System (INIS)

    Burtraw, D.; Harrison, K.; Pawlowski, J.A.

    1994-01-01

    Heavy trucks are primarily responsible for pavement damage to the nation's highways. In this paper we evaluate the pavement damage caused by coal trucks. We analyze the chief source of pavement damage (vehicle weight per axle, not total vehicle weight) and the chief cost involved (the periodic overlay that is required when a road's surface becomes worn). This analysis is presented in two stages. In the first section we present a synopsis of current economic theory including simple versions of the formulas that can be: used to calculate costs of pavement wear. In the second section we apply this theory to a specific example proximate to the reference environment for the Fuel Cycle Study in New Mexico in order to provide a numerical measure of the magnitude of the costs

  2. Natural resource damage assessment

    International Nuclear Information System (INIS)

    Seddelmeyer, J.

    1991-01-01

    The assessment and collection of natural resource damages from petroleum and chemical companies unfortunate enough to have injured publicly owned natural resources is perhaps the most rapidly expanding area of environmental liability. The idea of recovering for injury to publicly owned natural resources is an extension of traditional common law tort concepts under which a person who negligently injures another or his property is called upon to compensate the injured party. Normally, once liability has been established, it is a fairly straightforward matter to calculate the various elements of loss, such as the cost to repair or replace damaged property, or medical expenses, and lost income. More difficult questions, such as the amount to be awarded for pain and suffering or emotional distress, are left to the jury, although courts limit the circumstances in which the jury is permitted to award such damages

  3. Structural Basis of Mec1-Ddc2-RPA Assembly and Activation on Single-Stranded DNA at Sites of Damage.

    Science.gov (United States)

    Deshpande, Ishan; Seeber, Andrew; Shimada, Kenji; Keusch, Jeremy J; Gut, Heinz; Gasser, Susan M

    2017-10-19

    Mec1-Ddc2 (ATR-ATRIP) is a key DNA-damage-sensing kinase that is recruited through the single-stranded (ss) DNA-binding replication protein A (RPA) to initiate the DNA damage checkpoint response. Activation of ATR-ATRIP in the absence of DNA damage is lethal. Therefore, it is important that damage-specific recruitment precedes kinase activation, which is achieved at least in part by Mec1-Ddc2 homodimerization. Here, we report a structural, biochemical, and functional characterization of the yeast Mec1-Ddc2-RPA assembly. High-resolution co-crystal structures of Ddc2-Rfa1 and Ddc2-Rfa1-t11 (K45E mutant) N termini and of the Ddc2 coiled-coil domain (CCD) provide insight into Mec1-Ddc2 homodimerization and damage-site targeting. Based on our structural and functional findings, we present a Mec1-Ddc2-RPA-ssDNA composite structural model. By way of validation, we show that RPA-dependent recruitment of Mec1-Ddc2 is crucial for maintaining its homodimeric state at ssDNA and that Ddc2's recruitment domain and CCD are important for Mec1-dependent survival of UV-light-induced DNA damage. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Taxifolin enhances andrographolide-induced mitotic arrest and apoptosis in human prostate cancer cells via spindle assembly checkpoint activation.

    Directory of Open Access Journals (Sweden)

    Zhong Rong Zhang

    Full Text Available Andrographolide (Andro suppresses proliferation and triggers apoptosis in many types of cancer cells. Taxifolin (Taxi has been proposed to prevent cancer development similar to other dietary flavonoids. In the present study, the cytotoxic and apoptotic effects of the addition of Andro alone and Andro and Taxi together on human prostate carcinoma DU145 cells were assessed. Andro inhibited prostate cancer cell proliferation by mitotic arrest and activation of the intrinsic apoptotic pathway. Although the effect of Taxi alone on DU145 cell proliferation was not significant, the combined use of Taxi with Andro significantly potentiated the anti-proliferative effect of increased mitotic arrest and apoptosis by enhancing the cleavage of poly(ADP-ribose polymerase, and caspases-7 and -9. Andro together with Taxi enhanced microtubule polymerization in vitro, and they induced the formation of twisted and elongated spindles in the cancer cells, thus leading to mitotic arrest. In addition, we showed that depletion of MAD2, a component in the spindle assembly checkpoint (SAC, alleviated the mitotic block induced by the two compounds, suggesting that they trigger mitotic arrest by SAC activation. This study suggests that the anti-cancer activity of Andro can be significantly enhanced in combination with Taxi by disrupting microtubule dynamics and activating the SAC.

  5. Taxifolin Enhances Andrographolide-Induced Mitotic Arrest and Apoptosis in Human Prostate Cancer Cells via Spindle Assembly Checkpoint Activation

    Science.gov (United States)

    Wong, Matthew Man-Kin; Chiu, Sung-Kay; Cheung, Hon-Yeung

    2013-01-01

    Andrographolide (Andro) suppresses proliferation and triggers apoptosis in many types of cancer cells. Taxifolin (Taxi) has been proposed to prevent cancer development similar to other dietary flavonoids. In the present study, the cytotoxic and apoptotic effects of the addition of Andro alone and Andro and Taxi together on human prostate carcinoma DU145 cells were assessed. Andro inhibited prostate cancer cell proliferation by mitotic arrest and activation of the intrinsic apoptotic pathway. Although the effect of Taxi alone on DU145 cell proliferation was not significant, the combined use of Taxi with Andro significantly potentiated the anti-proliferative effect of increased mitotic arrest and apoptosis by enhancing the cleavage of poly(ADP-ribose) polymerase, and caspases-7 and -9. Andro together with Taxi enhanced microtubule polymerization in vitro, and they induced the formation of twisted and elongated spindles in the cancer cells, thus leading to mitotic arrest. In addition, we showed that depletion of MAD2, a component in the spindle assembly checkpoint (SAC), alleviated the mitotic block induced by the two compounds, suggesting that they trigger mitotic arrest by SAC activation. This study suggests that the anti-cancer activity of Andro can be significantly enhanced in combination with Taxi by disrupting microtubule dynamics and activating the SAC. PMID:23382917

  6. Zwint-1 is required for spindle assembly checkpoint function and kinetochore-microtubule attachment during oocyte meiosis.

    Science.gov (United States)

    Woo Seo, Dong; Yeop You, Seung; Chung, Woo-Jae; Cho, Dong-Hyung; Kim, Jae-Sung; Su Oh, Jeong

    2015-10-21

    The key step for faithful chromosome segregation during meiosis is kinetochore assembly. Defects in this process result in aneuploidy, leading to miscarriages, infertility and various birth defects. However, the roles of kinetochores in homologous chromosome segregation during meiosis are ill-defined. Here we found that Zwint-1 is required for homologous chromosome segregation during meiosis. Knockdown of Zwint-1 accelerated the first meiosis by abrogating the kinetochore recruitment of Mad2, leading to chromosome misalignment and a high incidence of aneuploidy. Although Zwint-1 knockdown did not affect Aurora C kinase activity, the meiotic defects following Zwint-1 knockdown were similar to those observed with ZM447439 treatment. Importantly, the chromosome misalignment following Aurora C kinase inhibition was not restored after removing the inhibitor in Zwint-1-knockdown oocytes, whereas the defect was rescued after the inhibitor washout in the control oocytes. These results suggest that Aurora C kinase-mediated correction of erroneous kinetochore-microtubule attachment is primarily regulated by Zwint-1. Our results provide the first evidence that Zwint-1 is required to correct erroneous kinetochore-microtubule attachment and regulate spindle checkpoint function during meiosis.

  7. Current Diagnosis and Management of Immune Related Adverse Events (irAEs Induced by Immune Checkpoint Inhibitor Therapy

    Directory of Open Access Journals (Sweden)

    Vivek Kumar

    2017-02-01

    Full Text Available The indications of immune checkpoint inhibitors (ICIs are set to rise further with the approval of newer agent like atezolimumab for use in patients with advanced stage urothelial carcinoma. More frequent use of ICIs has improved our understanding of their unique side effects, which are known as immune-related adverse events (irAEs. The spectrum of irAEs has expanded beyond more common manifestations such as dermatological, gastrointestinal and endocrine effects to rarer presentations involving nervous, hematopoietic and urinary systems. There are new safety data accumulating on ICIs in patients with previously diagnosed autoimmune conditions. It is challenging for clinicians to continuously update their working knowledge to diagnose and manage these events successfully. If diagnosed timely, the majority of events are completely reversible, and temporary immunosuppression with glucocorticoids, infliximab or other agents is warranted only in the most severe grade illnesses. The same principles of management will possibly apply as newer anti- cytotoxic T lymphocytes-associated antigen 4 (CTLA-4 and programmed cell death protein 1 (PD-1/PD-L1 antibodies are introduced. The current focus of research is for prophylaxis and for biomarkers to predict the onset of these toxicities. In this review we summarize the irAEs of ICIs and emphasize their growing spectrum and their management algorithms, to update oncology practitioners.

  8. The accuracy of survival time prediction for patients with glioma is improved by measuring mitotic spindle checkpoint gene expression.

    Directory of Open Access Journals (Sweden)

    Li Bie

    Full Text Available Identification of gene expression changes that improve prediction of survival time across all glioma grades would be clinically useful. Four Affymetrix GeneChip datasets from the literature, containing data from 771 glioma samples representing all WHO grades and eight normal brain samples, were used in an ANOVA model to screen for transcript changes that correlated with grade. Observations were confirmed and extended using qPCR assays on RNA derived from 38 additional glioma samples and eight normal samples for which survival data were available. RNA levels of eight major mitotic spindle assembly checkpoint (SAC genes (BUB1, BUB1B, BUB3, CENPE, MAD1L1, MAD2L1, CDC20, TTK significantly correlated with glioma grade and six also significantly correlated with survival time. In particular, the level of BUB1B expression was highly correlated with survival time (p<0.0001, and significantly outperformed all other measured parameters, including two standards; WHO grade and MIB-1 (Ki-67 labeling index. Measurement of the expression levels of a small set of SAC genes may complement histological grade and other clinical parameters for predicting survival time.

  9. The Spindle Assembly Checkpoint Is Not Essential for Viability of Human Cells with Genetically Lowered APC/C Activity.

    Science.gov (United States)

    Wild, Thomas; Larsen, Marie Sofie Yoo; Narita, Takeo; Schou, Julie; Nilsson, Jakob; Choudhary, Chunaram

    2016-03-01

    The anaphase-promoting complex/cyclosome (APC/C) and the spindle assembly checkpoint (SAC), which inhibits the APC/C, are essential determinants of mitotic timing and faithful division of genetic material. Activation of the APC/C is known to depend on two APC/C-interacting E2 ubiquitin-conjugating enzymes-UBE2C and UBE2S. We show that APC/C activity in human cells is tuned by the combinatorial use of three E2s, namely UBE2C, UBE2S, and UBE2D. Genetic deletion of UBE2C and UBE2S, individually or in combination, leads to discriminative reduction in APC/C function and sensitizes cells to UBE2D depletion. Reduction of APC/C activity results in loss of switch-like metaphase-to-anaphase transition and, strikingly, renders cells insensitive to chemical inhibition of MPS1 and genetic ablation of MAD2, both of which are essential for the SAC. These results provide insights into the regulation of APC/C activity and demonstrate that the essentiality of the SAC is imposed by the strength of the APC/C. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  10. The Spindle Assembly Checkpoint Is Not Essential for Viability of Human Cells with Genetically Lowered APC/C Activity

    DEFF Research Database (Denmark)

    Wild, Thomas; Larsen, Marie Sofie Yoo; Narita, Takeo

    2016-01-01

    The anaphase-promoting complex/cyclosome (APC/C) and the spindle assembly checkpoint (SAC), which inhibits the APC/C, are essential determinants of mitotic timing and faithful division of genetic material. Activation of the APC/C is known to depend on two APC/C-interacting E2 ubiquitin......-conjugating enzymes-UBE2C and UBE2S. We show that APC/C activity in human cells is tuned by the combinatorial use of three E2s, namely UBE2C, UBE2S, and UBE2D. Genetic deletion of UBE2C and UBE2S, individually or in combination, leads to discriminative reduction in APC/C function and sensitizes cells to UBE2D...... depletion. Reduction of APC/C activity results in loss of switch-like metaphase-to-anaphase transition and, strikingly, renders cells insensitive to chemical inhibition of MPS1 and genetic ablation of MAD2, both of which are essential for the SAC. These results provide insights into the regulation of APC...

  11. Immune Checkpoint Targets for Host-Directed Therapy to Prevent and Treat Leishmaniasis

    Directory of Open Access Journals (Sweden)

    Rajiv Kumar

    2017-11-01

    Full Text Available Leishmaniasis encompasses a group of diseases caused by protozoan parasites belonging to the genus Leishmania. These diseases range from life threatening visceral forms to self-healing cutaneous lesions, and each disease manifestations can progress to complications involving dissemination of parasites to skin or mucosal tissue. A feature of leishmaniasis is the key role host immune responses play in disease outcome. T cells are critical for controlling parasite growth. However, they can also contribute to disease onset and progression. For example, potent regulatory T cell responses can develop that suppress antiparasitic immunity. Alternatively, hyperactivated CD4+ or CD8+ T cells can be generated that cause damage to host tissues. There is no licensed human vaccine and drug treatment options are often limited and problematic. Hence, there is an urgent need for new strategies to improve the efficacy of current vaccine candidates and/or enhance both antiparasitic drug effectiveness and subsequent immunity in treated individuals. Here, we describe our current understanding about host immune responses contributing to disease protection and progression in the various forms of leishmaniasis. We also discuss how this knowledge may be used to develop new strategies for host-directed immune therapy to prevent or treat leishmaniasis. Given the major advances made in immune therapy in the cancer and autoimmune fields in recent years, there are significant opportunities to ride on the back of these successes in the infectious disease domain. Conversely, the rapid progress in our understanding about host immune responses during leishmaniasis is also providing opportunities to develop novel immunotherapy strategies that could have broad applications in diseases characterized by inflammation or immune dysfunction.

  12. [Transcranial magnetic stimulation and motor cortex stimulation in neuropathic pain].

    Science.gov (United States)

    Mylius, V; Ayache, S S; Teepker, M; Kappus, C; Kolodziej, M; Rosenow, F; Nimsky, C; Oertel, W H; Lefaucheur, J P

    2012-12-01

    Non-invasive and invasive cortical stimulation allows the modulation of therapy-refractory neuropathic pain. High-frequency repetitive transcranial magnetic stimulation (rTMS) of the contralateral motor cortex yields therapeutic effects at short-term and predicts the benefits of epidural motor cortex stimulation (MCS). The present article summarizes the findings on application, mechanisms and therapeutic effects of cortical stimulation in neuropathic pain.

  13. Grating stimulated echo

    International Nuclear Information System (INIS)

    Dubetsky, B.; Berman, P.R.; Sleator, T.

    1992-01-01

    A theory of a grating simulated echo (GTE) is developed. The GSE involves the sequential excitation of atoms by two counterpropagating traveling waves, a standing wave, and a third traveling wave. It is shown that the echo signal is very sensitive to small changes in atomic velocity, much more sensitive than the normal stimulated echo. Use of the GSE as a collisional probe or accelerometer is discussed

  14. Thyroid Stimulating Hormone Receptor

    Directory of Open Access Journals (Sweden)

    Murat Tuncel

    2017-02-01

    Full Text Available Thyroid stimulating hormone receptor (TSHR plays a pivotal role in thyroid hormone metabolism. It is a major controller of thyroid cell function and growth. Mutations in TSHR may lead to several thyroid diseases, most commonly hyperthyroidism. Although its genetic and epigenetic alterations do not directly lead to carcinogenesis, it has a crucial role in tumor growth, which is initiated by several oncogenes. This article will provide a brief review of TSHR and related diseases.

  15. mapDamage

    DEFF Research Database (Denmark)

    Ginolhac, Aurélien; Rasmussen, Morten; Gilbert, Tom

    2011-01-01

    Ancient DNA extracts consist of a mixture of contaminant DNA molecules, most often originating from environmental microbes, and endogenous fragments exhibiting substantial levels of DNA damage. The latter introduce specific nucleotide misincorporations and DNA fragmentation signatures in sequenci...... of the SAMtools suite and R environment and has been validated on both GNU/Linux and MacOSX operating systems....

  16. Core damage risk indicators

    International Nuclear Information System (INIS)

    Szikszai, T.

    1994-01-01

    The purpose of this document is to show a method for the fast recalculation of the PSA. To avoid the information loose, it is necessary to simplify the PSA models, or at least reorganize them. The method, introduced in this document, require that preparation, so we try to show, how to do that. This document is an introduction. This is the starting point of the work related to the development of the risk indicators. In the future, with the application of this method, we are going to show an everyday use of the PSA results to produce the indicators of the core damage risk. There are two different indicators of the plant safety performance, related to the core damage risk. The first is the core damage frequency indicator (CDFI), and the second is the core damage probability indicator (CDPI). Of course, we cannot describe all of the possible ways to use these indicators, rather we will try to introduce the requirements to establish such an indicator system and the calculation process

  17. Risk of nuclear damage

    International Nuclear Information System (INIS)

    Kienzl, K.

    1997-01-01

    Following the opening and words of welcome by Mr. Fritz Unterpertinger (unit director at the Austrian Federal Ministry for the Environment, Youth and Family; BMUJF) Mrs Helga Kromp-Kolb (professor at the Institute for Meteorology and Physics of the University of Natural Resources Science Vienna) illustrated the risks of nuclear damage in Europe by means of a nuclear risk map. She explained that even from a scientific or technical point of view the assessment of risks arising from nuclear power stations was fraught with great uncertainties. Estimates about in how far MCAs (maximum credible accident) could still be controlled by safety systems vary widely and so do assessments of the probability of a core melt. But there is wide agreement in all risk assessments conducted so far that MCAs might occur within a - from a human point of view - conceivable number of years. In this connection one has to bear in mind that the occurrence of such a major accident - whatever its probability may be - could entail immense damage and the question arises whether or not it is at all justifiable to expose the general public to such a risk. Klaus Rennings (Centre for European Economic Research, Mannheim, Germany) dealt with the economic aspects of nuclear risk assessment. He explained that there are already a number of studies available aiming to assess the risk of damage resulting from a core melt accident in economic terms. As to the probability of occurrence estimates vary widely between one incident in 3,333 and 250,000 year of reactor operation. It is assumed, however, that a nuclear accident involving a core melt in Germany would probably exceed the damage caused by the Chernobyl accident. The following speakers addressed the legal aspects of risks associated with nuclear installations. Mrs Monika Gimpel-Hinteregger (professor at the Institute for Civil Law in Graz) gave an overview on the applicable Austrian law concerning third party liability in the field of nuclear energy

  18. Disorder of G2-M Checkpoint Control in Aniline-Induced Cell Proliferation in Rat Spleen.

    Directory of Open Access Journals (Sweden)

    Jianling Wang

    Full Text Available Aniline, a toxic aromatic amine, is known to cause hemopoietic toxicity both in humans and animals. Aniline exposure also leads to toxic response in spleen which is characterized by splenomegaly, hyperplasia, fibrosis and the eventual formation of tumors on chronic in vivo exposure. Previously, we have shown that aniline exposure leads to iron overload, oxidative DNA damage, and increased cell proliferation, which could eventually contribute to a tumorigenic response in the spleen. Despite our demonstration that cell proliferation was associated with deregulation of G1 phase cyclins and increased expression of G1 phase cyclin-dependent kinases (CDKs, molecular mechanisms, especially the regulation of G2 phase and contribution of epigenetic mechanisms in aniline-induced splenic cellular proliferation remain largely unclear. This study therefore, mainly focused on the regulation of G2 phase in an animal model preceding a tumorigenic response. Male Sprague-Dawley rats were given aniline (0.5 mmol/kg/day in drinking water or drinking water only (controls for 30 days, and expression of G2 phase cyclins, CDK1, CDK inhibitors and miRNAs were measured in the spleen. Aniline treatment resulted in significant increases in cell cycle regulatory proteins, including cyclins A, B and CDK1, particularly phosphor-CDK1, and decreases in CDK inhibitors p21 and p27, which could promote the splenocytes to go through G2/M transition. Our data also showed upregulation of tumor markers Trx-1 and Ref-1 in rats treated with aniline. More importantly, we observed lower expression of miRNAs including Let-7a, miR-15b, miR24, miR-100 and miR-125, and greater expression of CDK inhibitor regulatory miRNAs such as miR-181a, miR-221 and miR-222 in the spleens of aniline-treated animals. Our findings suggest that significant increases in the expression of cyclins, CDK1 and aberrant regulation of miRNAs could lead to an accelerated G2/M transition of the splenocytes, and

  19. THE ATM-SMC1 PATHWAY IS ESSENTIAL FOR ACTIVATION OF THE CHROMIUM[VI]-INDUCED S-PHASE CHECKPOINT

    Science.gov (United States)

    Hexavalent chromium (Cr[VI] is a common industrial waste product, an environmental pollutant, and a recognized human carcinogen. Following cellular uptake, Cr[VI] can cause DNA damage, however, the mechanisims by which mammalian cells respond to Cr-induced DNA damage remain to b...

  20. Electrical Stimulation of Coleopteran Muscle for Initiating Flight.

    Directory of Open Access Journals (Sweden)

    Hao Yu Choo

    Full Text Available Some researchers have long been interested in reconstructing natural insects into steerable robots or vehicles. However, until recently, these so-called cyborg insects, biobots, or living machines existed only in science fiction. Owing to recent advances in nano/micro manufacturing, data processing, and anatomical and physiological biology, we can now stimulate living insects to induce user-desired motor actions and behaviors. To improve the practicality and applicability of airborne cyborg insects, a reliable and controllable flight initiation protocol is required. This study demonstrates an electrical stimulation protocol that initiates flight in a beetle (Mecynorrhina torquata, Coleoptera. A reliable stimulation protocol was determined by analyzing a pair of dorsal longitudinal muscles (DLMs, flight muscles that oscillate the wings. DLM stimulation has achieved with a high success rate (> 90%, rapid response time (< 1.0 s, and small variation (< 0.33 s; indicating little habituation. Notably, the stimulation of DLMs caused no crucial damage to the free flight ability. In contrast, stimulation of optic lobes, which was earlier demonstrated as a successful flight initiation protocol, destabilized the beetle in flight. Thus, DLM stimulation is a promising secure protocol for inducing flight in cyborg insects or biobots.

  1. Role of sound stimulation in reprogramming brain connectivity.

    Science.gov (United States)

    Chaudhury, Sraboni; Nag, Tapas C; Jain, Suman; Wadhwa, Shashi

    2013-09-01

    Sensory stimulation has a critical role to play in the development of an individual. Environmental factors tend to modify the inputs received by the sensory pathway. The developing brain is most vulnerable to these alterations and interacts with the environment to modify its neural circuitry. In addition to other sensory stimuli, auditory stimulation can also act as external stimuli to provide enrichment during the perinatal period. There is evidence that suggests that enriched environment in the form of auditory stimulation can play a substantial role in modulating plasticity during the prenatal period. This review focuses on the emerging role of prenatal auditory stimulation in the development of higher brain functions such as learning and memory in birds and mammals. The molecular mechanisms of various changes in the hippocampus following sound stimulation to effect neurogenesis, learning and memory are described. Sound stimulation can also modify neural connectivity in the early postnatal life to enhance higher cognitive function or even repair the secondary damages in various neurological and psychiatric disorders. Thus, it becomes imperative to examine in detail the possible ameliorating effects of prenatal sound stimulation in existing animal models of various psychiatric disorders, such as autism.

  2. Low intensity transcranial electric stimulation

    DEFF Research Database (Denmark)

    Antal, Andrea; Alekseichuk, I; Bikson, M

    2017-01-01

    Low intensity transcranial electrical stimulation (TES) in humans, encompassing transcranial direct current (tDCS), transcutaneous spinal Direct Current Stimulation (tsDCS), transcranial alternating current (tACS), and transcranial random noise (tRNS) stimulation or their combinations, appears...

  3. Human RECQL5beta stimulates flap endonuclease 1

    DEFF Research Database (Denmark)

    Speina, Elzbieta; Dawut, Lale; Hedayati, Mohammad

    2010-01-01

    devoid of RECQL1 and RECQL5 display increased chromosomal instability. Here, we report the physical and functional interaction of the large isomer of RECQL5, RECQL5beta, with the human flap endonuclease 1, FEN1, which plays a critical role in DNA replication, recombination and repair. RECQL5beta...... dramatically stimulates the rate of FEN1 cleavage of flap DNA substrates. Moreover, we show that RECQL5beta and FEN1 interact physically and co-localize in the nucleus in response to DNA damage. Our findings, together with the previous literature on WRN, BLM and RECQL4's stimulation of FEN1, suggests...

  4. Immunoassay of DNA damage

    International Nuclear Information System (INIS)

    Gasparro, F.P.; Santella, R.M.

    1988-01-01

    The direct photomodification of DNA by ultraviolet light or the photo-induced addition of exogenous compounds to DNA components results in alterations of DNA structure ranging from subtle to profound. There are two consequences of these conformational changes. First, cells in which the DNA has been damaged are capable of executing repair steps. Second, the DNA which is usually of very low immunogenicity now becomes highly antigenic. This latter property has allowed the production of a series of monoclonal antibodies that recognize photo-induced DNA damage. Monoclonal antibodies have been generated that recognize the 4',5'-monoadduct and the crosslink of 8-methoxypsoralen in DNA. In addition, another antibody has been prepared which recognizes the furan-side monoadduct of 6,4,4'-trimethylangelicin in DNA. These monoclonal antibodies have been characterized as to sensitivity and specificity using non-competitive and competitive enzyme-linked-immunosorbent assays (ELISA). (author)

  5. Neutron induced radiation damage

    International Nuclear Information System (INIS)

    Williams, M.M.R.

    1977-01-01

    We derive a general expression for the number of displaced atoms of type j caused by a primary knock-on of type i. The Kinchin-Pease model is used, but considerably generalised to allow for realistic atomic potentials. Two cases are considered in detail: the single particle problem causing a cascade and the neutron initiated problem which leads to multiple subcascades. Numerical results have been obtained for a variety of scattering laws. An important conclusion is that neutron initiated damage is much more severe than atom-initiated damage and leads to the number of displaced atoms being a factor of (A+1) 2 /4A larger than the single primary knock-on theory predicts. A is the ratio of the atomic mass to the neutron mass. The importance of this result to the theory of neutron sputtering is explained. (orig.) [de

  6. Immunoassay of DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Gasparro, F P; Santella, R M

    1988-09-01

    The direct photomodification of DNA by ultraviolet light or the photo-induced addition of exogenous compounds to DNA components results in alterations of DNA structure ranging from subtle to profound. There are two consequences of these conformational changes. First, cells in which the DNA has been damaged are capable of executing repair steps. Second, the DNA which is usually of very low immunogenicity now becomes highly antigenic. This latter property has allowed the production of a series of monoclonal antibodies that recognize photo-induced DNA damage. Monoclonal antibodies have been generated that recognize the 4',5'-monoadduct and the crosslink of 8-methoxypsoralen in DNA. In addition, another antibody has been prepared which recognizes the furan-side monoadduct of 6,4,4'-trimethylangelicin in DNA. These monoclonal antibodies have been characterized as to sensitivity and specificity using non-competitive and competitive enzyme-linked-immunosorbent assays (ELISA).

  7. Radiation damage in DNA

    International Nuclear Information System (INIS)

    Lafleur, V.

    1978-01-01

    A number of experiments are described with the purpose to obtain a better insight in the chemical nature and the biological significance of radiation-induced damage in DNA, with some emphasis on the significance of alkali-labile sites. It is shown that not only reactions of OH radicals but also of H radicals introduce breaks and other inactivating damage in single-standed phiX174 DNA. It is found that phosphate buffer is very suitable for the study of the reactions of H radicals with DNA, as the H 2 PO 4 - ions convert the hydrated electrons into H radicals. The hydrated electron, which does react with DNA, does not cause a detectable inactivation. (Auth.)

  8. Radiation damage of uranium

    International Nuclear Information System (INIS)

    Lazarevic, Dj.

    1966-11-01

    Study of radiation damage covered the following: Kinetics of electric resistance of uranium and uranium alloy with 1% of molybdenum dependent on the second phase and burnup rate; Study of gas precipitation and diffusion of bubbles by transmission electron microscopy; Numerical analysis of the influence of defects distribution and concentration on the rare gas precipitation in uranium; study of thermal sedimentation of uranium alloy with molybdenum; diffusion of rare gas in metal by gas chromatography method

  9. Cavitation damage of ceramics

    International Nuclear Information System (INIS)

    Kovalenko, V.I.; Marinin, V.G.

    1988-01-01

    Consideration is given to results of investigation of ceramic material damage under the effect of cavitation field on their surface, formed in water under the face of exponential concentrator, connected with ultrasonic generator UZY-3-0.4. Amplitude of vibrations of concentrator face (30+-2)x10 -6 m, frequency-21 kHz. It was established that ceramics resistance to cavitation effect correlated with the product of critical of stress intensity factor and material hardness

  10. Nondestructive damage detection and evaluation technique for seismically damaged structures

    Science.gov (United States)

    Adachi, Yukio; Unjoh, Shigeki; Kondoh, Masuo; Ohsumi, Michio

    1999-02-01

    The development of quantitative damage detection and evaluation technique, and damage detection technique for invisible damages of structures are required according to the lessons from the 1995 Hyogo-ken Nanbu earthquake. In this study, two quantitative damage sensing techniques for highway bridge structures are proposed. One method is to measure the change of vibration characteristics of the bridge structure. According to the damage detection test for damaged bridge column by shaking table test, this method can successfully detect the vibration characteristic change caused by damage progress due to increment excitations. The other method is to use self-diagnosis intelligent materials. According to the reinforced concrete beam specimen test, the second method can detect the damage by rupture of intelligent sensors, such as optical fiber or carbon fiber reinforced plastic rod.

  11. Spinal Cord Stimulation

    DEFF Research Database (Denmark)

    Meier, Kaare

    2014-01-01

    Spinal cord stimulation (SCS) is a surgical treatment for chronic neuropathic pain that is refractory to other treatment. Originally described by Shealy et al. in 1967(1), it is used to treat a range of conditions such as complex regional pain syndrome (CRPS I)(2), angina pectoris(3), radicular...... pain after failed back surgery syndrome (FBSS)(4), pain due to peripheral nerve injury, stump pain(5), peripheral vascular disease(6) and diabetic neuropathy(7,8); whereas phantom pain(9), postherpetic neuralgia(10), chronic visceral pain(11), and pain after partial spinal cord injury(12) remain more...

  12. Inflammation, oxidative DNA damage, and carcinogenesis

    International Nuclear Information System (INIS)

    Lewis, J.G.; Adams, D.O.

    1987-01-01

    Inflammation has long been associated with carcinogenesis, especially in the promotion phase. The mechanism of action of the potent inflammatory agent and skin promoter 12-tetradecanoyl phorbol-13-acetate (TPA) is unknown. It is though that TPA selectively enhances the growth of initiated cells, and during this process, initiated cells progress to the preneoplastic state and eventually to the malignant phenotype. The authors and others have proposed that TPA may work, in part, by inciting inflammation and stimulating inflammatory cells to release powerful oxidants which then induce DNA damage in epidermal cells. Macrophages cocultured with target cells and TPA induce oxidized thymine bases in the target cells. This process is inhibited by both catalase and inhibitors of lipoxygenases, suggesting the involvement of both H 2 O 2 and oxidized lipid products. In vivo studies demonstrated that SENCAR mice, which are sensitive to promotion by TPA, have a more intense inflammatory reaction in skin that C57LB/6 mice, which are resistant to promotion by TPA. In addition, macrophages from SENCAR mice release more H 2 O 2 and metabolites of AA, and induce more oxidative DNA damage in cocultured cells than macrophages from C57LB/6 mice. These data support the hypothesis that inflammation and the release of genotoxic oxidants may be one mechanism whereby initiated cells receive further genetic insults. They also further complicate risk assessment by suggesting that some environmental agents may work indirectly by subverting host systems to induce damage rather than maintaining homeostasis

  13. MO-FG-BRA-04: Leveraging the Abscopal Effect Via New Design Radiotherapy Biomaterials Loaded with Immune Checkpoint Inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Hao, Y; Cifter, G; Altundal, Y; Moreau, M; Sajo, E [Univ Massachusetts Lowell, Lowell, MA (United States); Sinha, N [Wentworth Institute of Technology, Boston, MA (United States); Makrigiorgos, G [Dana Farber Cancer Institute, Boston, MA (United States); Harvard Medical School, Boston, MA (United States); Ngwa, W [Univ Massachusetts Lowell, Lowell, MA (United States); Dana Farber Cancer Institute, Boston, MA (United States); Harvard Medical School, Boston, MA (United States)

    2015-06-15

    Purpose: Studies show that stereotactic body radiation therapy (SBRT) of a primary tumor in combination with immune checkpoint inhibitors (ICI) could Result in an immune-mediated regression of metastasis outside the radiation field, a phenomenon known as abscopal effect. However toxicities due to repeated systematic administration of ICI have been shown to be a major obstacle in clinical trials. Towards overcoming these toxicity limitations, we investigate a potential new approach whereby the ICI are administered via sustained in-situ release from radiotherapy (RT) biomaterials (e.g. fiducials) coated with a polymer containing the ICI. Methods: New design RT biomaterials were prepared by coating commercially available spacers/fiducials with a biocompatible polymer (PLGA) film containing fluorescent nanoparticles of size needed to load the ICI. The release of the nanoparticles was investigated in-vitro. Meanwhile, an experimentally determined in- vivo nanoparticle diffusion coefficient was employed in analytic calculations based on Fick’s second law to estimate the time for achieving the concentrations of ICI in the tumor draining lymph node (TDLN) that are needed to engender the abscopal effect during SBRT. The ICI investigated here was anti-CTLA-4 antibody (ipilimumab) at approved FDA concentrations. Results: Our in -vitro study results showed that RT biomaterials could be designed to achieve burst release of nanoparticles within one day. Meanwhile, our calculations indicate that for a 2 to 4 cm tumor it would take 4–22 days, respectively, following burst release, for the required concentration of ICI nanoparticles to accumulate in the TDLN during SBRT. Conclusion: Current investigations combining RT and immunotherapy involve repeated intravenous administration of ICI leading to significant systemic toxicities. Our preliminary results highlight a potential new approach for sustained in-situ release of the ICI from new design RT biomaterials. These results

  14. KinD is a checkpoint protein linking spore formation to extracellular-matrix production in Bacillus subtilis biofilms.

    Science.gov (United States)

    Aguilar, Claudio; Vlamakis, Hera; Guzman, Alejandra; Losick, Richard; Kolter, Roberto

    2010-05-18

    Bacillus subtilis cells form multicellular biofilm communities in which spatiotemporal regulation of gene expression occurs, leading to differentiation of multiple coexisting cell types. These cell types include matrix-producing and sporulating cells. Extracellular matrix production and sporulation are linked in that a mutant unable to produce matrix is delayed for sporulation. Here, we show that the delay in sporulation is not due to a growth advantage of the matrix-deficient mutant under these conditions. Instead, we show that the link between matrix production and sporulation is through the Spo0A signaling pathway. Both processes are regulated by the phosphorylated form of the master transcriptional regulator Spo0A. When cells have low levels of phosphorylated Spo0A (Spo0A~P), matrix genes are expressed; however, at higher levels of Spo0A~P, sporulation commences. We have found that Spo0A~P levels are maintained at low levels in the matrix-deficient mutant, thereby delaying expression of sporulation-specific genes. This is due to the activity of one of the components of the Spo0A phosphotransfer network, KinD. A deletion of kinD suppresses the sporulation defect of matrix mutants, while its overproduction delays sporulation. Our data indicate that KinD displays a dual role as a phosphatase or a kinase and that its activity is linked to the presence of extracellular matrix in the biofilms. We propose a novel role for KinD in biofilms as a checkpoint protein that regulates the onset of sporulation by inhibiting the activity of Spo0A until matrix, or a component therein, is sensed.

  15. Immune checkpoint inhibitor PD-1 pathway is down-regulated in synovium at various stages of rheumatoid arthritis disease progression.

    Science.gov (United States)

    Guo, Yanxia; Walsh, Alice M; Canavan, Mary; Wechalekar, Mihir D; Cole, Suzanne; Yin, Xuefeng; Scott, Brittney; Loza, Mathew; Orr, Carl; McGarry, Trudy; Bombardieri, Michele; Humby, Frances; Proudman, Susanna M; Pitzalis, Costantino; Smi