The Expression, Purification, and Characterization of a Ras Oncogene (Bras2) in Silkworm (Bombyx mori).

Science.gov (United States)

Lv, Zhengbing; Wang, Tao; Zhuang, Wenhua; Wang, Dan; Chen, Jian; Nie, Zuoming; Liu, Lili; Zhang, Wenping; Wang, Lisha; Wang, Deming; Wu, Xiangfu; Li, Jun; Qian, Lian; Zhang, Yaozhou

2013-01-01

The Ras oncogene of silkworm pupae (Bras2) may belong to the Ras superfamily. It shares 77% of its amino acid identity with teratocarcinoma oncogene 21 (TC21) related ras viral oncogene homolog-2 (R-Ras2) and possesses an identical core effector region. The mRNA of Bombyx mori Bras2 has 1412 bp. The open reading frame contains 603 bp, which encodes 200 amino acid residues. This recombinant BmBras2 protein was subsequently used as an antigen to raise a rabbit polyclonal antibody. Western blotting and real-time PCR analyses showed that BmBras2 was expressed during four developmental stages. The BmBras2 expression level was the highest in the pupae and was low in other life cycle stages. BmBras2 was expressed in all eight tested tissues, and it was highly expressed in the head, intestine, and epidermis. Subcellular localization studies indicated that BmBras2 was predominantly localized in the nuclei of Bm5 cells, although cytoplasmic staining was also observed to a lesser extent. A cell proliferation assay showed that rBmBras2 could stimulate the proliferation of hepatoma cells. The higher BmBras2 expression levels in the pupal stage, tissue expression patterns, and a cell proliferation assay indicated that BmBras2 promotes cell division and proliferation, most likely by influencing cell signal transduction.

Multiple fractions of gamma rays do not induce overexpression of c-myc or c-Ki-ras oncogenes in human cervical carcinoma cells

International Nuclear Information System (INIS)

Osmak, M.; Soric, J.; Matulic, M.

1993-01-01

Multiple fractions of gamma rays (0.5 Gy daily, 30 fractions) had previously been found to change the sensitivity of human cervical carcinoma HeLa cells to anticancer drugs. Preirradiated cells became resistant to cisplatin, methotrexate and vincristine but retained the same sensitivity to gamma rays and ultraviolet light. Some mechanisms involved in the resistance of preirradiated cells to cisplatin and vincristine were determined, i.e. the increased levels of metallothioneins and increased expression of plasma membrane P glycoprotein. As recent reports indicated that the resistance to cisplatin and ionizing radiation may involve the expression of oncogenes, the problem was studied whether multiple fractions of gamma rays can change the expression of c-myc and c-Ki-ras oncogenes in HeLa cells and whether there is a correlation between the expression of these oncogenes and the sensitivity of preirradiated cells to cisplatin and gamma rays. The expression of c-myc and c-Ki-ras oncogenes was examined using the DNA dot blot, the RNA dot blot and Northern blot analysis. The results show that preirradiation induced neither amplification nor elevated expression of c-myc and c-Ki-ras oncogenes. Furthermore, there is no correlation between the expression of c-myc and c-Ki-ras oncogenes and the acquired resistance to cisplatin. (author) 3 figs., 32 refs

A New Strategy to Control and Eradicate "Undruggable" Oncogenic K-RAS-Driven Pancreatic Cancer: Molecular Insights and Core Principles Learned from Developmental and Evolutionary Biology.

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Van Sciver, Robert E; Lee, Michael P; Lee, Caroline Dasom; Lafever, Alex C; Svyatova, Elizaveta; Kanda, Kevin; Colliver, Amber L; Siewertsz van Reesema, Lauren L; Tang-Tan, Angela M; Zheleva, Vasilena; Bwayi, Monicah N; Bian, Minglei; Schmidt, Rebecca L; Matrisian, Lynn M; Petersen, Gloria M; Tang, Amy H

2018-05-14

Oncogenic K-RAS mutations are found in virtually all pancreatic cancers, making K-RAS one of the most targeted oncoproteins for drug development in cancer therapies. Despite intense research efforts over the past three decades, oncogenic K-RAS has remained largely "undruggable". Rather than targeting an upstream component of the RAS signaling pathway (i.e., EGFR/HER2) and/or the midstream effector kinases (i.e., RAF/MEK/ERK/PI3K/mTOR), we propose an alternative strategy to control oncogenic K-RAS signal by targeting its most downstream signaling module, Seven-In-Absentia Homolog (SIAH). SIAH E3 ligase controls the signal output of oncogenic K-RAS hyperactivation that drives unchecked cell proliferation, uncontrolled tumor growth, and rapid cancer cell dissemination in human pancreatic cancer. Therefore, SIAH is an ideal therapeutic target as it is an extraordinarily conserved downstream signaling gatekeeper indispensable for proper RAS signaling. Guided by molecular insights and core principles obtained from developmental and evolutionary biology, we propose an anti-SIAH-centered anti-K-RAS strategy as a logical and alternative anticancer strategy to dampen uncontrolled K-RAS hyperactivation and halt tumor growth and metastasis in pancreatic cancer. The clinical utility of developing SIAH as both a tumor-specific and therapy-responsive biomarker, as well as a viable anti-K-RAS drug target, is logically simple and conceptually innovative. SIAH clearly constitutes a major tumor vulnerability and K-RAS signaling bottleneck in pancreatic ductal adenocarcinoma (PDAC). Given the high degree of evolutionary conservation in the K-RAS/SIAH signaling pathway, an anti-SIAH-based anti-PDAC therapy will synergize with covalent K-RAS inhibitors and direct K-RAS targeted initiatives to control and eradicate pancreatic cancer in the future.

Biological aspects and tumorigenic activity of the Ras proto-oncogenic family Aspectos biológicos e atividade tumorigênica da família proto-oncogênica Ras

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Juliano André Boquett

2010-12-01

Full Text Available Proto-oncogenes play an important role in the regulation of the cellular cycle, being critical to the tumorigenesis. In this category we can find the RAS family. Due to the high transformation potential of these genes, this family is the best described and most studied one. It is formed by the H-, K- and the N-RAS genes, that codify highly related proteins expressed in several types of cells, denominated p21.These proteins act in the sign transduction from the membrane to the nucleus, as well as in the control of proliferation, differentiation and cellular death, and they are regulated by the interaction with GDP (inactive and GTP (active. These proteins show variation in only 10 - 15% of the primary structure, in the C-terminal portion denominated hyper-variant region. When in the oncogenic form, the p21 proteins remain active, providing continuous stimuli to the cellular proliferation. Among the RAS genes, K-RAS ones have been the most studied for presenting more frequent mutations and for being present in more aggressive tumors, implying the patients’ shorter survival time. Due to these facts and relative bibliography lack in the Portuguese language on this family, we presented in this work a systematized and updated review on the RAS genes. Os proto-oncogenes desempenham importante papel na regulação do ciclo celular, e são críticos à tumorigênese. Nessa categoria se encontra a família RAS, que, devido ao elevado potencial transformante dos genes que a compõem, é uma das mais bem descritas e estudadas. É formada pelos genes H-, K- e N-RAS, que codificam proteínas altamente relacionadas expressas em vários tipos de células, denominadas p21. Estas atuam na transdução de sinal da membrana ao núcleo, estão envolvidas no controle da proliferação, diferenciação e morte celular e são reguladas pela interação com GDP (inativa e GTP (ativa. As proteínas p21 diferem em apenas 10-15% da sua estrutura primária, na porção C

Senescence-Associated Secretory Phenotypes Reveal Cell-Nonautonomous Functions of Oncogenic RAS and the p53 Tumor Suppressor

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Copp& #233; , Jean-Philippe; Patil, Christopher; Rodier, Francis; Sun, Yu; Munoz, Denise; Goldstein, Joshua; Nelson, Peter; Desprez, Pierre-Yves; Campisi, Judith

2008-10-24

Cellular senescence suppresses cancer by arresting cell proliferation, essentially permanently, in response to oncogenic stimuli, including genotoxic stress. We modified the use of antibody arrays to provide a quantitative assessment of factors secreted by senescent cells. We show that human cells induced to senesce by genotoxic stress secrete myriad factors associated with inflammation and malignancy. This senescence-associated secretory phenotype (SASP) developed slowly over several days and only after DNA damage of sufficient magnitude to induce senescence. Remarkably similar SASPs developed in normal fibroblasts, normal epithelial cells, and epithelial tumor cells after genotoxic stress in culture, and in epithelial tumor cells in vivo after treatment of prostate cancer patients with DNA-damaging chemotherapy. In cultured premalignant epithelial cells, SASPs induced an epithelial-mesenchyme transition and invasiveness, hallmarks of malignancy, by a paracrine mechanism that depended largely on the SASP factors interleukin (IL)-6 and IL-8. Strikingly, two manipulations markedly amplified, and accelerated development of, the SASPs: oncogenic RAS expression, which causes genotoxic stress and senescence in normal cells, and functional loss of the p53 tumor suppressor protein. Both loss of p53 and gain of oncogenic RAS also exacerbated the promalignant paracrine activities of the SASPs. Our findings define a central feature of genotoxic stress-induced senescence. Moreover, they suggest a cell-nonautonomous mechanism by which p53 can restrain, and oncogenic RAS can promote, the development of age-related cancer by altering the tissue microenvironment.

Stat1 phosphorylation determines Ras oncogenicity by regulating p27 kip1.

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Shuo Wang

Full Text Available Inactivation of p27 Kip1 is implicated in tumorigenesis and has both prognostic and treatment-predictive values for many types of human cancer. The transcription factor Stat1 is essential for innate immunity and tumor immunosurveillance through its ability to act downstream of interferons. Herein, we demonstrate that Stat1 functions as a suppressor of Ras transformation independently of an interferon response. Inhibition of Ras transformation and tumorigenesis requires the phosphorylation of Stat1 at tyrosine 701 but is independent of Stat1 phosphorylation at serine 727. Stat1 induces p27 Kip1 expression in Ras transformed cells at the transcriptional level through mechanisms that depend on Stat1 phosphorylation at tyrosine 701 and activation of Stat3. The tumor suppressor properties of Stat1 in Ras transformation are reversed by the inactivation of p27 Kip1. Our work reveals a novel functional link between Stat1 and p27 Kip1, which act in coordination to suppress the oncogenic properties of activated Ras. It also supports the notion that evaluation of Stat1 phosphorylation in human tumors may prove a reliable prognostic factor for patient outcome and a predictor of treatment response to anticancer therapies aimed at activating Stat1 and its downstream effectors.

Quantitative two-dimensional gel electrophoresis analysis of human fibroblasts transformed by ras oncogenes.

Science.gov (United States)

Miller, M J; Maher, V M; McCormick, J J

1992-11-01

Quantitative two-dimensional gel electrophoresis was used to compare the cellular protein patterns of a normal foreskin-derived human fibroblasts cell line (LG1) and three immortal derivatives of LG1. One derivative, designated MSU-1.1 VO, was selected for its ability to grow in the absence of serum and is non-tumorigenic in athymic mice. The other two strains were selected for focus-formation following transfection with either Ha-ras or N-ras oncogenes and form high grade malignant tumors. Correspondence and cluster analysis provided a nonbiased estimate of the relative similarity of the different two-dimensional patterns. These techniques separated the gel patterns into three distinct classes: LG1, MSU-1.1 VO, and the ras transformed cell strains. The MSU-1.1 VO cells were more closely related to the parental LG1 than to the ras-transformed cells. The differences between the three classes were primarily quantitative in nature: 16% of the spots demonstrated statistically significant changes (P 2) in the rate of incorporation of radioactive amino acids. The patterns from the two ras-transformed cell strains were similar, and variations in the expression of proteins that occurred between the separate experiments obscured consistent differences between the Ha-ras and N-ras transformed cells. However, while only 9 out of 758 spots were classified as different (1%), correspondence analysis could consistently separate the two ras transformants. One of these spots was five times more intense in the Ha-ras transformed cells than the N-ras.(ABSTRACT TRUNCATED AT 250 WORDS)

Ha-ras oncogene expression directed by a milk protein gene promoter: tissue specificity, hormonal regulation, and tumor induction in transgenic mice

International Nuclear Information System (INIS)

Andres, A.C.; Schoenenberger, C.A.; Groner, B.; Henninghausen, L.; LeMeur, M.; Gelinger, P.

1987-01-01

The activated human Ha-ras oncogene was subjected to the control of the promoter region of the murine whey acidic protein (Wap) gene, which is expressed in mammary epithelial cells in response to lactogenic hormones. The Wap-ras gene was stably introduced into the mouse germ line of five transgenic mice (one male and four females). Wap-ras expression was observed in the mammary glands of lactating females in two lines derived from female founders. The tissue-directed and hormone-dependent Wap expression was conferred on the Ha-ras oncogene. The signals governing Wap expression are located within 2.5 kilobases of 5' flanking sequence. The other two lines derived from female founders did not express the chimeric gene. In the line derived from the male founder the Wap-ras gene is integrated into the Y chromosome. Expression was found in the salivary gland of male animals only. After a long latency, Wap-ras-expressing mice developed tumors. The tumors arose in tissues expressing Wap-ras - i.e., mammary or salivary glands. Compared to the corresponding nonmalignant tissues, Wap-ras expression was enhanced in the tumors

SCD1 Expression is dispensable for hepatocarcinogenesis induced by AKT and Ras oncogenes in mice.

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Lei Li

Full Text Available Increased de novo lipogenesis is one of the major metabolic events in cancer. In human hepatocellular carcinoma (HCC, de novo lipogenesis has been found to be increased and associated with the activation of AKT/mTOR signaling. In mice, overexpression of an activated form of AKT results in increased lipogenesis and hepatic steatosis, ultimately leading to liver tumor development. Hepatocarcinogenesis is dramatically accelerated when AKT is co-expressed with an oncogenic form of N-Ras. SCD1, the major isoform of stearoyl-CoA desaturases, catalyzing the conversion of saturated fatty acids (SFA into monounsaturated fatty acids (MUFA, is a key enzyme involved in de novo lipogenesis. While many studies demonstrated the requirement of SCD1 for tumor cell growth in vitro, whether SCD1 is necessary for tumor development in vivo has not been previously investigated. Here, we show that genetic ablation of SCD1 neither inhibits lipogenesis and hepatic steatosis in AKT-overexpressing mice nor affects liver tumor development in mice co-expressing AKT and Ras oncogenes. Molecular analysis showed that SCD2 was strongly upregulated in liver tumors from AKT/Ras injected SCD1(-/- mice. Noticeably, concomitant silencing of SCD1 and SCD2 genes was highly detrimental for the growth of AKT/Ras cells in vitro. Altogether, our study provides the evidence, for the first time, that SCD1 expression is dispensable for AKT/mTOR-dependent hepatic steatosis and AKT/Ras-induced hepatocarcinogenesis in mice. Complete inhibition of stearoyl-CoA desaturase activity may be required to efficiently suppress liver tumor development.

The human T-lymphotropic virus type I tax gene can cooperate with the ras oncogene to induce neoplastic transformation of cells.

Science.gov (United States)

Pozzatti, R; Vogel, J; Jay, G

1990-01-01

Epidemiologic studies have linked infection by the human T-lymphotropic virus type I (HTLV-I) with the development of adult T-cell leukemia. The low penetrance of the virus and the long latency for disease manifestation are factors that obscure the role of HTLV-I infection in oncogenesis. We have used an in vitro transformation assay system to determine directly whether the HTLV-I tax gene has transformation potential. Transfection of the tax gene alone into early-passage rat embryo fibroblasts did not induce morphological alterations. However, cotransfection of tax with the selectable marker plasmid pRSVneo gave rise to G418-resistant colonies that could be established as immortalized cell lines. Cotransfection of tax with the ras oncogene into rat embryo fibroblasts gave rise to foci of transformed cells that were highly tumorigenic in nude mice. These data represent a direct demonstration of the oncogenic potential of the tax gene in nonlymphoid cells and establish HTLV-I as a transforming virus.

Expression of ras oncogene and major histocompatibility complex (MHC) antigen in carcinomas of the uterine cervix

International Nuclear Information System (INIS)

Cho, Kyung Ja; Jang, Ja June; Kim, Yong Dae; Ha, Chang Won; Koh, Jae Soo

1993-01-01

Consecutive 50 cases of squamous cell carcinomas of the uterine cervix diagnosed in 1992 were subjected to immunohistochemical study for ras oncogene product (p21) and MHC class II (DR) antigen using a microprobe immunostainer. Activated ras and aberrant DR expression were noted in 26 cases (52%) and 11 cases (22%) of cervical squamous cell carcinomas, respectively, without difference among histologic types. The reaction was mainly intracytoplasmic, with granular staining pattern and diffuse distribution. No direct histologic correlation between ras and DR expression was found. Four cases with HPV 16/18 DNA in superficial koilocytotic cells, revealed by in situ hybridization, showed various expression of ras and DR, and these 3 factors histologically did not seem to be affected one another. (Author)

GTP Binding and Oncogenic Mutations May Attenuate Hypervariable Region (HVR)-Catalytic Domain Interactions in Small GTPase K-Ras4B, Exposing the Effector Binding Site*

Science.gov (United States)

Lu, Shaoyong; Banerjee, Avik; Jang, Hyunbum; Zhang, Jian; Gaponenko, Vadim; Nussinov, Ruth

2015-01-01

K-Ras4B, a frequently mutated oncogene in cancer, plays an essential role in cell growth, differentiation, and survival. Its C-terminal membrane-associated hypervariable region (HVR) is required for full biological activity. In the active GTP-bound state, the HVR interacts with acidic plasma membrane (PM) headgroups, whereas the farnesyl anchors in the membrane; in the inactive GDP-bound state, the HVR may interact with both the PM and the catalytic domain at the effector binding region, obstructing signaling and nucleotide exchange. Here, using molecular dynamics simulations and NMR, we aim to figure out the effects of nucleotides (GTP and GDP) and frequent (G12C, G12D, G12V, G13D, and Q61H) and infrequent (E37K and R164Q) oncogenic mutations on full-length K-Ras4B. The mutations are away from or directly at the HVR switch I/effector binding site. Our results suggest that full-length wild-type GDP-bound K-Ras4B (K-Ras4BWT-GDP) is in an intrinsically autoinhibited state via tight HVR-catalytic domain interactions. The looser association in K-Ras4BWT-GTP may release the HVR. Some of the oncogenic mutations weaken the HVR-catalytic domain association in the K-Ras4B-GDP/-GTP bound states, which may facilitate the HVR disassociation in a nucleotide-independent manner, thereby up-regulating oncogenic Ras signaling. Thus, our results suggest that mutations can exert their effects in more than one way, abolishing GTP hydrolysis and facilitating effector binding. PMID:26453300

C/EBPβ represses p53 to promote cell survival downstream of DNA damage independent of oncogenic Ras and p19Arf

Science.gov (United States)

Ewing, SJ; Zhu, S; Zhu, F; House, JS; Smart, RC

2013-01-01

CCAAT/enhancer-binding protein-β (C/EBPβ) is a mediator of cell survival and tumorigenesis. When C/EBPβ−/− mice are treated with carcinogens that produce oncogenic Ras mutations in keratinocytes, they respond with abnormally elevated keratinocyte apoptosis and a block in skin tumorigenesis. Although this aberrant carcinogen-induced apoptosis results from abnormal upregulation of p53, it is not known whether upregulated p53 results from oncogenic Ras and its ability to induce p19Arf and/or activate DNA-damage response pathways or from direct carcinogen-induced DNA damage. We report that p19Arf is dramatically elevated in C/EBPβ−/− epidermis and that C/EBPβ represses a p19Arf promoter reporter. To determine whether p19Arf is responsible for the proapoptotic phenotype in C/EBPβ−/− mice, C/EBPβ−/−;p19Arf−/− mice were generated. C/EBPβ−/−;p19Arf−/− mice responded to carcinogen treatment with increased p53 and apoptosis, indicating p19Arf is not essential. To ascertain whether oncogenic Ras activation induces aberrant p53 and apoptosis in C/EBPβ−/− epidermis, we generated K14-ER:Ras; C/EBPβ−/− mice. Oncogenic Ras activation induced by 4-hydroxytamoxifen did not produce increased p53 or apoptosis. Finally, when C/EBPβ−/− mice were treated with differing types of DNA-damaging agents, including alkylating chemotherapeutic agents, they displayed aberrant levels of p53 and apoptosis. These results indicate that C/EBPβ represses p53 to promote cell survival downstream of DNA damage and suggest that inhibition of C/EBPβ may be a target for cancer cotherapy to increase the efficacy of alkylating chemotherapeutic agents. PMID:18636078

GTP Binding and Oncogenic Mutations May Attenuate Hypervariable Region (HVR)-Catalytic Domain Interactions in Small GTPase K-Ras4B, Exposing the Effector Binding Site.

Science.gov (United States)

Lu, Shaoyong; Banerjee, Avik; Jang, Hyunbum; Zhang, Jian; Gaponenko, Vadim; Nussinov, Ruth

2015-11-27

K-Ras4B, a frequently mutated oncogene in cancer, plays an essential role in cell growth, differentiation, and survival. Its C-terminal membrane-associated hypervariable region (HVR) is required for full biological activity. In the active GTP-bound state, the HVR interacts with acidic plasma membrane (PM) headgroups, whereas the farnesyl anchors in the membrane; in the inactive GDP-bound state, the HVR may interact with both the PM and the catalytic domain at the effector binding region, obstructing signaling and nucleotide exchange. Here, using molecular dynamics simulations and NMR, we aim to figure out the effects of nucleotides (GTP and GDP) and frequent (G12C, G12D, G12V, G13D, and Q61H) and infrequent (E37K and R164Q) oncogenic mutations on full-length K-Ras4B. The mutations are away from or directly at the HVR switch I/effector binding site. Our results suggest that full-length wild-type GDP-bound K-Ras4B (K-Ras4B(WT)-GDP) is in an intrinsically autoinhibited state via tight HVR-catalytic domain interactions. The looser association in K-Ras4B(WT)-GTP may release the HVR. Some of the oncogenic mutations weaken the HVR-catalytic domain association in the K-Ras4B-GDP/-GTP bound states, which may facilitate the HVR disassociation in a nucleotide-independent manner, thereby up-regulating oncogenic Ras signaling. Thus, our results suggest that mutations can exert their effects in more than one way, abolishing GTP hydrolysis and facilitating effector binding. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Characterization of a human MSX-2 cDNA and its fragment isolated as a transformation suppressor gene against v-Ki-ras oncogene.

Science.gov (United States)

Takahashi, C; Akiyama, N; Matsuzaki, T; Takai, S; Kitayama, H; Noda, M

1996-05-16

A cDNA (termed CT124) encoding a carboxyl-terminal fragment of the human homeobox protein MSX-2 was found to induce flat reversion when expressed in v-Ki-ras-transformed NIH3T3 cells. Although the expression of endogenous MSX-2 gene is low in most of the normal adult tissues examined, it is frequently activated in carcinoma-derived cell lines. Likewise, the gene is inactive in NIH3T3 cells but is transcriptionally activated after transformation by v-Ki-ras oncogene, suggesting that the intact MSX-2 may play a positive, rather than suppressive, role in cell transformation. To test this possibility, we isolated a near full-length human MSX-2 cDNA and tested its activities in two cell systems, i.e. fibroblast and myoblast. In NIH3T3 fibroblasts, although the gene by itself failed to confer a transformed phenotype, antisense MSX-2 cDNA as well as truncated CT124 cDNA interfered with the transforming activities of v-Ki-ras oncogene. In C2C12 myoblasts, MSX-2 was found to suppress MyoD gene expression, as do activated ras oncogenes, under certain culture conditions, and CT124 was found to inhibit the activities of both MSX-2 and ras in this system as well. Our findings not only suggest that CT124 may act as a dominant suppressor of MSX-2 but also raise the possibility that MSX-2 gene may be an important downstream target for the Ras signaling pathways.

H-Ras and K-Ras Oncoproteins Induce Different Tumor Spectra When Driven by the Same Regulatory Sequences.

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Drosten, Matthias; Simón-Carrasco, Lucía; Hernández-Porras, Isabel; Lechuga, Carmen G; Blasco, María T; Jacob, Harrys K C; Fabbiano, Salvatore; Potenza, Nicoletta; Bustelo, Xosé R; Guerra, Carmen; Barbacid, Mariano

2017-02-01

Genetic studies in mice have provided evidence that H-Ras and K-Ras proteins are bioequivalent. However, human tumors display marked differences in the association of RAS oncogenes with tumor type. Thus, to further assess the bioequivalence of oncogenic H-Ras and K-Ras, we replaced the coding region of the murine K-Ras locus with H-Ras G12V oncogene sequences. Germline expression of H-Ras G12V or K-Ras G12V from the K-Ras locus resulted in embryonic lethality. However, expression of these genes in adult mice led to different tumor phenotypes. Whereas H-Ras G12V elicited papillomas and hematopoietic tumors, K-Ras G12V induced lung tumors and gastric lesions. Pulmonary expression of H-Ras G12V created a senescence-like state caused by excessive MAPK signaling. Likewise, H-Ras G12V but not K-Ras G12V induced senescence in mouse embryonic fibroblasts. Label-free quantitative analysis revealed that minor differences in H-Ras G12V expression levels led to drastically different biological outputs, suggesting that subtle differences in MAPK signaling confer nonequivalent functions that influence tumor spectra induced by RAS oncoproteins. Cancer Res; 77(3); 707-18. ©2016 AACR. ©2016 American Association for Cancer Research.

Transformation by Oncogenic Ras Expands the Early Genomic Response to Transforming Growth Factor β in Intestinal Epithelial Cells

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Carl E. Allen

2008-10-01

Full Text Available A substantial body of evidence implicates TGFβ as a tumor promoter in epithelial cells that have become resistant to its tumor suppressor activity. To better understand early, genome-wide TGFβ responses in cells resistant to growth inhibition by TGFβ, we used microarray analysis in a well-defined cell culture system of sensitive and resistant intestinal epithelial cells. TGFβ-regulated gene expression in TGFβ-growth-sensitive, nontransformed rat intestinal epithelial cells (RIE-1 was compared to expression in TGFβ-growth-resistant RIE cells stably transformed by oncogenic Ras(12V. Treatment of RIE-1 cells with 2 ng/ml TGFβ1 for 1 hour increased the expression of eight gene sequences by 2.6-fold or more, whereas eight were down regulated 2.6-fold. In RIE-Ras(12V cells, 42 gene sequences were upregulated and only 3 were down-regulated. Comparison of RIE and RIE-Ras(12V identified 37 gene sequences as unique, Ras-dependent genomic targets of TGFβ1. TGFβ-regulation of connective tissue growth factor and vascular endothelial growth factor, two genes up-regulated in RIE-Ras cells and previously implicated in tumor promotion, was independently confirmed and further characterized by Northern analysis. Our data indicate that overexpression of oncogenic Ras in intestinal epithelial cells confers a significantly expanded repertoire of robust, early transcriptional responses to TGFβ via signaling pathways yet to be fully elucidated but including the canonical Raf-1/MAPK/Erk pathway. Loss of sensitivity to growth inhibition by TGFβ does not abrogate TGFβ signaling and actually expands the early transcriptional response to TGFβ1. Expression of some of these genes may confer to Ras-transformed cells characteristics favorable for tumor promotion.

Effect of ionizing radiation on the biological activity of activated oncogenes and dormant proto-oncogenes

International Nuclear Information System (INIS)

Angenent, G.C.; Berg, K.J. van den.

1984-01-01

The authors have studied the effect of ionizing radiation on the cloned human activated Ha-ras oncogene, on the Ha-ras gene in integrated form and on the dormant proto-oncogene murine c-mos using the NIH/3T3 transfection system. NIH/3T3 cells were transfected with DNA from the plasmid pT24 carrying the cloned Ha-ras oncogene of the T24 bladder carcinoma cell line. Various individual foci which developed were injected into nude mice. DNA was isolated from tumours, digested with the restriction enzyme Bam HI, electrophoresed on agarose and blotted onto nitrocellulose filter according to Southern. Hybridization with a pT24 probe showed that all the primary foci of transformed cells contained various fragments of the pT24 plasmid indicating that fibroblast transformation had been induced by introduction of the Ha-ras oncogene. (Auth.)

Concurrent mutation in exons 1 and 2 of the K-ras oncogene in colorectal cancer

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Fiorella Guadagni

2012-01-01

Full Text Available The K-ras gene is frequently mutated in colorectal cancer and has been associated with tumor initiation and progression; approximately 90% of the activating mutations are found in codons 12 and 13 of exon 1 and just under 5% in codon 61 located in exon 2. These mutations determine single aminoacidic substitutions in the GTPase pocket leading to a block of the GTP hydrolytic activity of the K-ras p21 protein, and therefore to its constitutive activation. Point mutations in sites of the K-ras gene, other than codons 12, 13 and 61, and other types of genetic alterations, may occur in a minority of cases, such as in the less frequent cases of double mutations in the K-ras gene. However, all mutations in this gene, even those which occur in non-canonical sites or double mutations, are relevant oncogenic alterations in colorectal cancer and may underlie K-ras pathway hyperactivation. In the present study, we report the case of a patient with colorectal cancer presenting a concurrent point mutation in exons 1 and 2 of the K-ras gene, a GGT to TGT substitution (Glycine to Cysteine at codon 12, and a GAC to AAC substitution (Aspartic Acid to Asparagine at codon 57. In addition, we found in the same patient’s sample a silent polymorphism at codon 11 (Ala11Ala of exon 1. (Folia Histochemica et Cytobiologica 2011; Vol. 49, No. 4, pp. 729–733

Transformation and radiosensitivity of human diploid skin fibroblasts transfected with activated RAS oncogene and SV40 T-antigen

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Su, L.-N.; Little, J.B. (Harvard School of Public Health, Boston, MA (United States))

1992-08-01

Three normal human diploid cell strains were transfected with an activated Ha-ras oncogene (EJ ras) or SV40 T-antigen. Multiple clones were examined for morphological alterations, growth requirements, ability to grow under anchorage independent conditions, immortality and tumorigenicity in nude mice. Clones expressing SV40 T-antigen alone or in combination with ras protein p21 were significantly radioresistant as compared with their parent cells or clones transfected with the neo gene only. This radioresistant phenotype persisted in post-crisis, immortalized cell lines. These data suggest that expression of the SV40 T-antigen but not activated Ha-ras plays an important role in the radiosensitivity of human diploid cells. The radioresistant phenotype in SV40 T transfected cells was not related to the enhanced level of genetic instability seen in pre-crisis and newly immortalized cells, nor to the process of immortalization itself. (author).

Transformation and radiosensitivity of human diploid skin fibroblasts transfected with activated RAS oncogene and SV40 T-antigen

International Nuclear Information System (INIS)

Su, L.-N.; Little, J.B.

1992-01-01

Three normal human diploid cell strains were transfected with an activated Ha-ras oncogene (EJ ras) or SV40 T-antigen. Multiple clones were examined for morphological alterations, growth requirements, ability to grow under anchorage independent conditions, immortality and tumorigenicity in nude mice. Clones expressing SV40 T-antigen alone or in combination with ras protein p21 were significantly radioresistant as compared with their parent cells or clones transfected with the neo gene only. This radioresistant phenotype persisted in post-crisis, immortalized cell lines. These data suggest that expression of the SV40 T-antigen but not activated Ha-ras plays an important role in the radiosensitivity of human diploid cells. The radioresistant phenotype in SV40 T transfected cells was not related to the enhanced level of genetic instability seen in pre-crisis and newly immortalized cells, nor to the process of immortalization itself. (author)

The bovine papillomavirus E5 oncogene can cooperate with ras: identification of p21 amino acids critical for transformation by c-rasH but not v-rasH

DEFF Research Database (Denmark)

Willumsen, B M; Vass, W C; Velu, T J

1991-01-01

We have previously used a series of insertion-deletion mutants of the mutationally activated v-rasH gene to identify several regions of the encoded protein that are dispensable for cellular transformation (B. M. Willumsen, A. G. Papageorge, H.-F. Kung, E. Bekesi, T. Robins, M. Johnsen, W. C. Vass...... in their v-rasH forms. We conclude that a region including amino acids 102 and 103 encodes a function that is more critical to c-rasH than to v-rasH. Guanine nucleotide exchange is one function that is compatible with such a phenotype......., and D. R. Lowy, Mol. Cell. Biol. 6:2646-2654, 1986). To determine if some of these amino acids are more important for the biological activity of c-rasH, we have now tested many of the same insertion-deletion mutants in the c-rasH form for their ability to transform NIH 3T3 cells. Since the transforming...

Identification of a c-Jun N-terminal kinase-2-dependent signal amplification cascade that regulates c-Myc levels in ras transformation

DEFF Research Database (Denmark)

Mathiasen, D.P.; Egebjerg, C.; Andersen, S.H.

2012-01-01

are essential for ras transformation. Previous studies show that ERK-mediated serine 62 phosphorylation protects c-Myc from proteasomal degradation. ERK is, however, not alone sufficient to stabilize c-Myc but requires the cooperation of cancerous inhibitor of protein phosphatase 2A (CIP2A), an oncogene...... that counteracts protein phosphatase 2A-mediated dephosphorylation of c-Myc. Here we show that JNK2 regulates Cip2a transcription via ATF2. ATF2 and c-Myc cooperate to activate the transcription of ATF3. Remarkably, not only ectopic JNK2, but also ectopic ATF2, CIP2A, c-Myc and ATF3 are sufficient to rescue...... the defective ras transformation of JNK2-deficient cells. Thus, these data identify the key signal converging point of JNK2 and ERK pathways and underline the central role of CIP2A in ras transformation.Oncogene advance online publication, 27 June 2011; doi:10.1038/onc.2011.230....

Mitotic Stress Is an Integral Part of the Oncogene-Induced Senescence Program that Promotes Multinucleation and Cell Cycle Arrest

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Dina Dikovskaya

2015-09-01

Full Text Available Oncogene-induced senescence (OIS is a tumor suppression mechanism that blocks cell proliferation in response to oncogenic signaling. OIS is frequently accompanied by multinucleation; however, the origin of this is unknown. Here, we show that multinucleate OIS cells originate mostly from failed mitosis. Prior to senescence, mutant H-RasV12 activation in primary human fibroblasts compromised mitosis, concordant with abnormal expression of mitotic genes functionally linked to the observed mitotic spindle and chromatin defects. Simultaneously, H-RasV12 activation enhanced survival of cells with damaged mitoses, culminating in extended mitotic arrest and aberrant exit from mitosis via mitotic slippage. ERK-dependent transcriptional upregulation of Mcl1 was, at least in part, responsible for enhanced survival and slippage of cells with mitotic defects. Importantly, mitotic slippage and oncogene signaling cooperatively induced senescence and key senescence effectors p21 and p16. In summary, activated Ras coordinately triggers mitotic disruption and enhanced cell survival to promote formation of multinucleate senescent cells.

The inflammatory cytokine TNFα cooperates with Ras in elevating metastasis and turns WT-Ras to a tumor-promoting entity in MCF-7 cells

International Nuclear Information System (INIS)

Leibovich-Rivkin, Tal; Liubomirski, Yulia; Meshel, Tsipi; Abashidze, Anastasia; Brisker, Daphna; Solomon, Hilla; Rotter, Varda; Weil, Miguel; Ben-Baruch, Adit

2014-01-01

In the present study we determined the relative contribution of two processes to breast cancer progression: (1) Intrinsic events, such as activation of the Ras pathway and down-regulation of p53; (2) The inflammatory cytokines TNFα and IL-1β, shown in our published studies to be highly expressed in tumors of >80% of breast cancer patients with recurrent disease. Using MCF-7 human breast tumor cells originally expressing WT-Ras and WT-p53, we determined the impact of the above-mentioned elements and cooperativity between them on the expression of CXCL8 (ELISA, qRT-PCR), a member of a “cancer-related chemokine cluster” that we have previously identified. Then, we determined the mechanisms involved (Ras-binding-domain assays, Western blot, luciferase), and tested the impact of Ras + TNFα on angiogenicity (chorioallantoic membrane assays) and on tumor growth at the mammary fat pad of mice and on metastasis, in vivo. Using Ras G12V that recapitulates multiple stimulations induced by receptor tyrosine kinases, we found that Ras G12V alone induced CXCL8 expression at the mRNA and protein levels, whereas down-regulation of p53 did not. TNFα and IL-1β potently induced CXCL8 expression and synergized with Ras G12V , together leading to amplified CXCL8 expression. Testing the impact of WT-Ras, which is the common form in breast cancer patients, we found that WT-Ras was not active in promoting CXCL8; however, TNFα has induced the activation of WT-Ras: joining these two elements has led to cooperative induction of CXCL8 expression, via the activation of MEK, NF-κB and AP-1. Importantly, TNFα has led to increased expression of WT-Ras in an active GTP-bound form, with properties similar to those of Ras G12V . Jointly, TNFα + Ras activities have given rise to increased angiogenesis and to elevated tumor cell dissemination to lymph nodes. TNFα cooperates with Ras in promoting the metastatic phenotype of MCF-7 breast tumor cells, and turns WT-Ras into a tumor

Transformation and radiosensitivity of human diploid skin fibroblasts transfected with activated ras oncogene and SV40 T-antigen.

Science.gov (United States)

Su, L N; Little, J B

1992-08-01

Three normal human diploid cell strains were transfected with an activated Ha-ras oncogene (EJ ras) or SV40 T-antigen. Multiple clones were examined for morphological alterations, growth requirements, ability to grow under anchorage independent conditions, immortality and tumorigenicity in nude mice. Clones expressing SV40 T-antigen alone or in combination with ras protein p21 were significantly radioresistant as compared with their parent cells or clones transfected with the neo gene only. This radioresistant phenotype persisted in post-crisis, immortalized cell lines. Cells transfected with EJ ras alone showed no morphological alterations nor significant changes in radiosensitivity. Cell clones expressing ras and/or SV40 T-antigen showed a reduced requirement for serum supplements, an increase in aneuploidy and chromosomal aberrations, and enhanced growth in soft agar as an early cellular response to SV40 T-antigen expression. The sequential order of transfection with SV40 T-antigen and ras influenced radio-sensitivity but not the induction of morphological changes. These data suggest that expression of the SV40 T-antigen but not activated Ha-ras plays an important role in the radiosensitivity of human diploid cells. The radioresistant phenotype in SV40 T transfected cells was not related to the enhanced level of genetic instability seen in pre-crisis and newly immortalized cells, nor to the process of immortalization itself.

In vitro modeling of human pancreatic duct epithelial cell transformation defines gene expression changes induced by K-ras oncogenic activation in pancreatic carcinogenesis.

Science.gov (United States)

Qian, Jiaying; Niu, Jiangong; Li, Ming; Chiao, Paul J; Tsao, Ming-Sound

2005-06-15

Genetic analysis of pancreatic ductal adenocarcinomas and their putative precursor lesions, pancreatic intraepithelial neoplasias (PanIN), has shown a multistep molecular paradigm for duct cell carcinogenesis. Mutational activation or inactivation of the K-ras, p16(INK4A), Smad4, and p53 genes occur at progressive and high frequencies in these lesions. Oncogenic activation of the K-ras gene occurs in >90% of pancreatic ductal carcinoma and is found early in the PanIN-carcinoma sequence, but its functional roles remain poorly understood. We show here that the expression of K-ras(G12V) oncogene in a near diploid HPV16-E6E7 gene immortalized human pancreatic duct epithelial cell line originally derived from normal pancreas induced the formation of carcinoma in 50% of severe combined immunodeficient mice implanted with these cells. A tumor cell line established from one of these tumors formed ductal cancer when implanted orthotopically. These cells also showed increased activation of the mitogen-activated protein kinase, AKT, and nuclear factor-kappaB pathways. Microarray expression profiling studies identified 584 genes whose expression seemed specifically up-regulated by the K-ras oncogene expression. Forty-two of these genes have been reported previously as differentially overexpressed in pancreatic cancer cell lines or primary tumors. Real-time PCR confirmed the overexpression of a large number of these genes. Immunohistochemistry done on tissue microarrays constructed from PanIN and pancreatic cancer samples showed laminin beta3 overexpression starting in high-grade PanINs and occurring in >90% of pancreatic ductal carcinoma. The in vitro modeling of human pancreatic duct epithelial cell transformation may provide mechanistic insights on gene expression changes that occur during multistage pancreatic duct cell carcinogenesis.

Characterization of sur-2, a Novel Ras-Mediated Signal Transduction Component in C. elegans

National Research Council Canada - National Science Library

DesJardins, Edward

1998-01-01

... (oncogenes). A subset of proto-oncogenes comprise the RAS signal transduction pathway. Vulval development in the nematode worm Caenorhabditis elegans is controlled by a RAS signal transduction pathway...

Oncogenic Activation of Fibroblast Growth Factor Receptor-3 and RAS Genes as Non-Overlapping Mutual Exclusive Events in Urinary Bladder Cancer.

Science.gov (United States)

Pandith, Arshad A; Hussain, Aashaq; Khan, Mosin S; Shah, Zafar A; Wani, M Saleem; Siddiqi, Mushtaq A

2016-01-01

Urinary bladder cancer is a common malignancy in the West and ranks as the 7th most common cancer in our region of Kashmir, India. FGFR3 mutations are frequent in superficial urothelial carcinoma (UC) differing from the RAS gene mutational pattern. The aim of this study was to analyze the frequency and association of FGFR3 and RAS gene mutations in UC cases. Paired tumor and adjacent normal tissue specimens of 65 consecutive UC patients were examined. DNA preparations were evaluated for the occurrence of FGFR3 and RAS gene mutations by PCR-SCCP and DNA sequencing. Somatic point mutations of FGFR3 were identified in 32.3% (21 of 65). The pattern and distribution were significantly associated with low grade/stage (<0.05). The overall mutations in exon 1 and 2 in all the forms of RAS genes aggregated to 21.5% and showed no association with any clinic-pathological parameters. In total, 53.8% (35 of 65) of the tumors studied had mutations in either a RAS or FGFR3 gene, but these were totally mutually exclusive in and none of the samples showed both the mutational events in mutually exclusive RAS and FGFR3. We conclude that RAS and FGFR3 mutations in UC are mutually exclusive and non-overlapping events which reflect activation of oncogenic pathways through different elements.

Characterization of sur-2, a Novel Ras-Mediated Signal Transduction Component in C. elegans

National Research Council Canada - National Science Library

DesJardins, Edward

1999-01-01

... (oncogenes). A subset of proto-oncogenes comprise the RAS signal transduction pathway. Vulval development in the nematode worm Caenorhabditis elegans is controlled by a RAS signal transduction pathway. C...

Inhibition of carcinogen induced c-Ha-ras and c-fos proto-oncogenes expression by dietary curcumin

Directory of Open Access Journals (Sweden)

Lipigorngoson Suwiwek

2001-01-01

Full Text Available Abstract Background We investigated the chemopreventive action of dietary curcumin on 7,12-dimethylbenz(aanthracene (DMBA-initiated and 12,0-tetradecanoylphorbol-13-acetate (TPA-promoted skin tumor formation in Swiss albino mice. Curcumin, a yellow coloring matter isolated from roots of Curcuma longa Linn, is a phenolic compound possessing antioxidant, free radical scavenger, and antiinflammatory properties. It has been shown by previously reported work that TPA-induced skin tumors were inhibited by topical application of curcumin, and curcumin has been shown to inhibit a variety of biological activities of TPA. Topical application of curcumin was reported to inhibit TPA-induced c-fos, c-jun and c-myc gene expression in mouse skin. This paper reports the effects of orally administered curcumin, which was consumed as a dietary component at concentrations of 0.2 % or 1 %, in ad libitum feeding. Results Animals in which tumors had been initiated with DMBA and promoted with TPA experienced significantly fewer tumors and less tumor volume if they ingested either 0.2% or 1% curcumin diets. Also, the dietary consumption of curcumin resulted in a significantly decreased expression of ras and fos proto-oncogenes in the tumorous skin, as measured by enhanced chemiluminesence Western blotting detection system (Amersham. Conclusions Whereas earlier work demonstrated that topical application of curcumin to mouse skin inhibited TPA-induced expression of c-fos, c-jun and c-myc oncogenes, our results are the first to show that orally consumed curcumin significantly inhibited DMBA- and TPA-induced ras and fos gene expression in mouse skin.

Inhibition of carcinogen induced c-Ha-ras and c-fos proto-oncogenes expression by dietary curcumin

International Nuclear Information System (INIS)

Limtrakul, Porn-ngarm; Anuchapreeda, Songyot; Lipigorngoson, Suwiwek; Dunn, Floyd W

2001-01-01

We investigated the chemopreventive action of dietary curcumin on 7,12-dimethylbenz(a)anthracene (DMBA)-initiated and 12,0-tetradecanoylphorbol-13-acetate (TPA)-promoted skin tumor formation in Swiss albino mice. Curcumin, a yellow coloring matter isolated from roots of Curcuma longa Linn, is a phenolic compound possessing antioxidant, free radical scavenger, and antiinflammatory properties. It has been shown by previously reported work that TPA-induced skin tumors were inhibited by topical application of curcumin, and curcumin has been shown to inhibit a variety of biological activities of TPA. Topical application of curcumin was reported to inhibit TPA-induced c-fos, c-jun and c-myc gene expression in mouse skin. This paper reports the effects of orally administered curcumin, which was consumed as a dietary component at concentrations of 0.2 % or 1 %, in ad libitum feeding. Animals in which tumors had been initiated with DMBA and promoted with TPA experienced significantly fewer tumors and less tumor volume if they ingested either 0.2% or 1% curcumin diets. Also, the dietary consumption of curcumin resulted in a significantly decreased expression of ras and fos proto-oncogenes in the tumorous skin, as measured by enhanced chemiluminesence Western blotting detection system (Amersham). Whereas earlier work demonstrated that topical application of curcumin to mouse skin inhibited TPA-induced expression of c-fos, c-jun and c-myc oncogenes, our results are the first to show that orally consumed curcumin significantly inhibited DMBA- and TPA-induced ras and fos gene expression in mouse skin

INDUCTION OF DNA ADDUCTS, TUMORS, AND KI-RAS ONCOGENE MUTATIONS IN STRAIN A/J MOUSE LUNG BY IP. ADMINISTRATION OF DIBENZ[A,H]ANTHRACENE

Science.gov (United States)

Induction of DNA adducts, tumors, and Ki-ras oncogene mutations in strain AlJ mouse lung by ip. administration of dibenz[a,h]anthracene Previous studies of polycyclic aromatic hydrocarbon (P AH) induced lung tumors in the strain NJ mouse model system have demonstrated qua...

K-Ras(V14I) -induced Noonan syndrome predisposes to tumour development in mice.

Science.gov (United States)

Hernández-Porras, Isabel; Schuhmacher, Alberto J; Garcia-Medina, Raquel; Jiménez, Beatriz; Cañamero, Marta; de Martino, Alba; Guerra, Carmen

2016-06-01

The Noonan syndrome (NS) is an autosomal dominant genetic disorder characterized by short stature, craniofacial dysmorphism, and congenital heart defects. A significant proportion of NS patients may also develop myeloproliferative disorders (MPDs), including juvenile myelomonocytic leukaemia (JMML). Surprisingly, scarce information is available in relation to other tumour types in these patients. We have previously developed and characterized a knock-in mouse model that carries one of the most frequent KRAS-NS-related mutations, the K-Ras(V14I) substitution, which recapitulates most of the alterations described in NS patients, including MPDs. The K-Ras(V14I) mutation is a mild activating K-Ras protein; thus, we have used this model to study tumour susceptibility in comparison with mice expressing the classical K-Ras(G12V) oncogene. Interestingly, our studies have shown that these mice display a generalized tumour predisposition and not just MPDs. In fact, we have observed that the K-Ras(V14I) mutation is capable of cooperating with the p16Ink4a/p19Arf and Trp53 tumour suppressors, as well as with other risk factors such as pancreatitis, thereby leading to a higher cancer incidence. In conclusion, our results illustrate that the K-Ras(V14I) activating protein is able to induce cancer, although at a much lower level than the classical K-Ras(G12V) oncogene, and that it can be significantly modulated by both genetic and non-genetic events. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Oncogenic Ras-Induced Morphologic Change Is through MEK/ERK Signaling Pathway to Downregulate Stat3 at a Posttranslational Level in NIH3T3 Cells

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Hsuan-Heng Yeh

2008-01-01

Full Text Available Ras is a key regulator of the MAP kinase-signaling cascade and may cause morphologic change of Ras-transformed cells. Signal transducer and activator of transcription 3 (Stat3 can be activated by cytokine stimulation. In this study, we unravel that Ha-rasV12 overexpression can downregulate the expression of Stat3 protein at a posttranslational level in NIH3T3 cells. Furthermore, we demonstrate that Stat3 expression downregulated by Ha-rasV12 overexpression is through proteosome degradation and not through a mTOR/p70S6K-related signaling pathway. The suppression of Stat3 accompanied by the morphologic change induced by Ha-rasV12 was through mitogen extracellular kinase (MEK/extracellular-regulated kinase (ERK signaling pathway. Microtubule disruption is involved in Ha-rasV12-induced morphologic change, which could be reversed by overexpression of Stat3. Taken together, we are the first to demonstrate that Stat3 protein plays a critical role in Ha-rasV12-induced morphologic change. Oncogenic Ras-triggered morphologic change is through the activation of MEK/ERK to posttranslationally downregulate Stat3 expression. Our finding may shed light on developing novel therapeutic strategies against Ras-related tumorigenesis.

Rat embryo cells immortalized with transfected oncogenes are transformed by gamma irradiation.

Science.gov (United States)

Endlich, B; Salavati, R; Sullivan, T; Ling, C C

1992-12-01

Cesium-137 gamma rays were used to transform rat embryo cells (REC) which were first transfected with activated c-myc or c-Ha-ras oncogenes to produce immortal cell lines (REC:myc and REC:ras). When exposed to 6 Gy of 137Cs gamma rays, some cells became morphologically transformed with focus formation frequencies of approximately 3 x 10(-4) for REC:myc and approximately 1 x 10(-4) for REC:ras, respectively. Cells isolated from foci of gamma-ray-transformed REC:myc (REC:myc:gamma) formed anchorage-independent colonies and were tumorigenic in nude mice, but foci from gamma-ray-transformed REC:ras (REC:ras:gamma) did not exhibit either of these criteria of transformation. Similar to the results with gamma irradiation, we observed a sequence-dependent phenomenon when myc and ras were transfected into REC, one at a time. REC immortalized by ras transfection were not converted to a tumorigenic phenotype by secondary transfection with myc, but REC transfected with myc were very susceptible to transformation by subsequent ras transfection. This suggests that myc-immortalized cells are more permissive to transformation via secondary treatments. In sequentially transfected REC, myc expression was high whether it was transfected first or second, whereas ras expression was highest when the ras gene was transfected secondarily into myc-containing REC. Molecular analysis of REC:ras:gamma transformants showed no alterations in structure of the transfected ras or of the endogenous ras, myc, p53, or fos genes. The expression of ras and p53 was increased in some isolates of REC:ras:gamma, but myc and fos expression were not affected. Similarly, REC:myc:gamma transformants did not demonstrate rearrangement or amplification of the transfected or the endogenous myc genes, or of the potentially cooperating Ha-, Ki-, or N-ras genes. Northern hybridization analysis revealed increased expression of N-ras in two isolates, REC:myc:gamma 33 and gamma 41, but no alterations in the expression

Plac8 Links Oncogenic Mutations to Regulation of Autophagy and Is Critical to Pancreatic Cancer Progression

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Conan Kinsey

2014-05-01

Full Text Available Mutations in p53 and RAS potently cooperate in oncogenic transformation, and correspondingly, these genetic alterations frequently coexist in pancreatic ductal adenocarcinoma (PDA and other human cancers. Previously, we identified a set of genes synergistically activated by combined RAS and p53 mutations as frequent downstream mediators of tumorigenesis. Here, we show that the synergistically activated gene Plac8 is critical for pancreatic cancer growth. Silencing of Plac8 in cell lines suppresses tumor formation by blocking autophagy, a process essential for maintaining metabolic homeostasis in PDA, and genetic inactivation in an engineered mouse model inhibits PDA progression. We show that Plac8 is a critical regulator of the autophagic machinery, localizing to the lysosomal compartment and facilitating lysosome-autophagosome fusion. Plac8 thus provides a mechanistic link between primary oncogenic mutations and the induction of autophagy, a central mechanism of metabolic reprogramming, during PDA progression.

New insights into RAS biology reinvigorate interest in mathematical modeling of RAS signaling.

Science.gov (United States)

Erickson, Keesha E; Rukhlenko, Oleksii S; Posner, Richard G; Hlavacek, William S; Kholodenko, Boris N

2018-03-05

RAS is the most frequently mutated gene across human cancers, but developing inhibitors of mutant RAS has proven to be challenging. Given the difficulties of targeting RAS directly, drugs that impact the other components of pathways where mutant RAS operates may potentially be effective. However, the system-level features, including different localizations of RAS isoforms, competition between downstream effectors, and interlocking feedback and feed-forward loops, must be understood to fully grasp the opportunities and limitations of inhibiting specific targets. Mathematical modeling can help us discern the system-level impacts of these features in normal and cancer cells. New technologies enable the acquisition of experimental data that will facilitate development of realistic models of oncogenic RAS behavior. In light of the wealth of empirical data accumulated over decades of study and the advancement of experimental methods for gathering new data, modelers now have the opportunity to advance progress toward realization of targeted treatment for mutant RAS-driven cancers. Copyright © 2018 Elsevier Ltd. All rights reserved.

Mevalonates, Ras and Breast Cancer

National Research Council Canada - National Science Library

White, Michael

2001-01-01

.... This selective inhibition appears to be a consequence of expression of oncogenic Ras. Here we are evaluating the ability of Fmev to selectively interfere with proliferation of breast cancer cells...

RAS/ERK modulates TGFbeta-regulated PTEN expression in human pancreatic adenocarcinoma cells.

Science.gov (United States)

Chow, Jimmy Y C; Quach, Khai T; Cabrera, Betty L; Cabral, Jennifer A; Beck, Stayce E; Carethers, John M

2007-11-01

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is rarely mutated in pancreatic cancers, but its regulation by transforming growth factor (TGF)-beta might mediate growth suppression and other oncogenic actions. Here, we examined the role of TGFbeta and the effects of oncogenic K-RAS/ERK upon PTEN expression in the absence of SMAD4. We utilized two SMAD4-null pancreatic cell lines, CAPAN-1 (K-RAS mutant) and BxPc-3 (WT-K-RAS), both of which express TGFbeta surface receptors. Cells were treated with TGFbeta1 and separated into cytosolic/nuclear fractions for western blotting with phospho-SMAD2, SMAD 2, 4 phospho-ATP-dependent tyrosine kinases (Akt), Akt and PTEN antibodies. PTEN mRNA levels were assessed by reverse transcriptase-polymerase chain reaction. The MEK1 inhibitor, PD98059, was used to block the downstream action of oncogenic K-RAS/ERK, as was a dominant-negative (DN) K-RAS construct. TGFbeta increased phospho-SMAD2 in both cytosolic and nuclear fractions. PD98059 treatment further increased phospho-SMAD2 in the nucleus of both pancreatic cell lines, and DN-K-RAS further improved SMAD translocation in K-RAS mutant CAPAN cells. TGFbeta treatment significantly suppressed PTEN protein levels concomitant with activation of Akt by 48 h through transcriptional reduction of PTEN mRNA that was evident by 6 h. TGFbeta-induced PTEN suppression was reversed by PD98059 and DN-K-RAS compared with treatments without TGFbeta. TGFbeta-induced PTEN expression was inversely related to cellular proliferation. Thus, oncogenic K-RAS/ERK in pancreatic adenocarcinoma facilitates TGFbeta-induced transcriptional down-regulation of the tumor suppressor PTEN in a SMAD4-independent manner and could constitute a signaling switch mechanism from growth suppression to growth promotion in pancreatic cancers.

RAS/ERK modulates TGFβ-regulated PTEN expression in human pancreatic adenocarcinoma cells

OpenAIRE

Chow, Jimmy Y.C.; Quach, Khai T.; Cabrera, Betty L.; Cabral, Jennifer A.; Beck, Stayce E.; Carethers, John M.

2007-01-01

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is rarely mutated in pancreatic cancers, but its regulation by transforming growth factor (TGF)-β might mediate growth suppression and other oncogenic actions. Here, we examined the role of TGFβ and the effects of oncogenic K-RAS/ERK upon PTEN expression in the absence of SMAD4. We utilized two SMAD4-null pancreatic cell lines, CAPAN-1 (K-RAS mutant) and BxPc-3 (WT-K-RAS), both of which express TGFβ surface receptors. Cells were t...

Exploiting the bad eating habits of Ras-driven cancers.

Science.gov (United States)

White, Eileen

2013-10-01

Oncogenic Ras promotes glucose fermentation and glutamine use to supply central carbon metabolism, but how and why have only emerged recently. Ras-mediated metabolic reprogramming generates building blocks for growth and promotes antioxidant defense. To fuel metabolic pathways, Ras scavenges extracellular proteins and lipids. To bolster metabolism and mitigate stress, Ras activates cellular self-cannibalization and recycling of proteins and organelles by autophagy. Targeting these distinct features of Ras-driven cancers provides novel approaches to cancer therapy.

Diet, Lifestyle and risk of K-ras mutation-positive and -negative colorectal adenomas

NARCIS (Netherlands)

Wark, P.A.; Kuil, van der W.; Ploemacher, J.; Muijen, van G.N.P.; Mulder, Ch.J.J.; Weijenberg, M.P.; Kok, F.J.; Kampman, E.

2006-01-01

K-ras mutation-positive (K-ras+) and -negative (K-ras-) colorectal adenomas may differ clinically and pathologically. As environmental compounds may cause mutations in the growth-related K-ras oncogene or affect clonal selection depending on mutational status, we evaluated whether the aetiology of

Influence of a ras oncogene on platelet-derived growth factor (PDGF)-stimulated phosphoinositide hydrolysis in murine fibroblasts

International Nuclear Information System (INIS)

Parries, G.; Racker, E.

1986-01-01

The authors have examined the effects of transfection of rat-1 fibroblasts with the ras oncogene on the metabolism of phosphatidylinositol (PI). Incubation of [ 3 H]inositol-labeled rat-1 cells with PDGF resulted in a 2- to 3-fold increase in [ 3 H]IP3 levels within 90 s. In the presence of 25 mM Li+, [ 3 H]IP1 levels were increased 8-fold after 30 min. In contrast, incubation of ras-transfected fibroblasts (EJ-2 line) with PDGF had little or no effect on the level of either [ 3 H]IP3 or [ 3 H]IP1. Similar stimulations by PDGF were observed in NIH 3T3 cells, but not in Kirsten virus-transformed or Harvey ras-transfected cell lines. On the other hand, NIH 3T3 cells transfected with v-src responded to PDGF by stimulation of PI turnover similar to the parent cell line. In NIH 3T3 cells transfected with an expression vector containing the v-Ha-ras gene under transcriptional control of the glucocorticoid-inducible mouse mammary tumor virus promoter, the PDGF stimulation of [ 3 H]inositol incorporation into PI was reduced from 10-fold in the absence of dexamethasone to 1.8-fold when the cells were pretreated for 26 h with 2 μM dexamethasone. In the parental 3T3 cells PDGF stimulation was reduced by about 40% in the presence of dexamethasone. In the absence of PDGF the rate of PI turnover (i.e., the kinetics of [ 3 H]IP1 accumulation in the presence of Li+) in EJ-2 cells was similar to that in rat-1 cells. Thus, in the presence of PDGF, the rate of PI turnover in rat-1 cells was several fold higher than in the transfected cells. These results suggest that the ras gene product (p21) may exert an inhibitory effect on PDGF-stimulated phosphoinositide metabolism

Oncogenes and radiation resistance - a review

International Nuclear Information System (INIS)

Dritschilo, A.

1992-01-01

Oncogenes exert their effects on the genetic programs of cells by regulating signal transduction pathways, resulting in multi-factorial genetic responses. By such actions, the genetic elements responsible for the cellular responses to ionizing radiation may be affected. Reports implicating the association of oncogene expression with modulation of the radiation response include the ras, raf, and myc genes. Experiments overexpressing H-ras and c-raf-1 using genetically engineered constructs result in enhanced post-radiation cellular survival. Conversely, inhibition of raf gene expression has resulted in relative radiation sensitization and delay of human squamous cell carcinoma tumor growth in nude mice. There appears to be a potential strategy for therapeutic intervention. The identification of genes that confer survival advantage following radiation exposure, and understanding their mechanisms of action, may permit a genetically based intervention for radiation sensitization. One such approach employs oligo-deoxynucleotides complementary to oncogene-encoded in RNA's (antisense DNA). (author)

K-RAS and N-RAS mutations in testicular germ cell tumors

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Bekir Muhammet Hacioglu

2017-05-01

Full Text Available Testicular cancer is a relatively rare tumor type, accounting for approximately 1% of all cancers in men. However, among men aged between 15 and 40 years, testicular cancer is the most commonly diagnosed malignancy. Testicular germ cell tumors (TGCTs are classified as seminoma and non-seminoma. The RAS oncogene controls several cellular functions, including cell proliferation, apoptosis, migration, and differentiation. Thus, RAS signaling is important for normal germ cell development. Mutations of the Kirsten RAS (K-RAS gene are present in over 20% of all cancers. RAS gene mutations have also been reported in TGCTs. We investigated K-RAS and N-RAS mutations in seminoma and non-seminoma TGCT patients. A total of 24 (55% pure seminoma cases and 19 (45% non-seminoma cases were included in the study. K-RAS and N-RAS analyses were performed in our molecular pathology laboratory, using K-RAS and N-RAS Pyro Kit 24 V1 (Qiagen. In total, a RAS mutation was present in 12 patients (27%: 7 seminoma (29% and 5 non-seminoma cases (26% [p = 0.55]. A K-RAS mutation was present in 4 pure seminoma tumors (16% and 3 non-seminoma tumors (15% [p = 0.63], and an N-RAS mutation was observed in 4 seminoma tumors (16% and 3 non-seminoma tumors (15% [p = 0.63]. Both, K-RAS and N-RAS mutations were present in two patients: one with seminoma tumor and the other with non-seminoma tumor. To date, no approved targeted therapy is available for the treatment of TGCTs. The analysis of K-RAS and N-RAS mutations in these tumors may provide more treatment options, especially in platinum-resistant tumors.

ARF and ATM/ATR cooperate in p53-mediated apoptosis upon oncogenic stress

International Nuclear Information System (INIS)

Pauklin, Siim; Kristjuhan, Arnold; Maimets, Toivo; Jaks, Viljar

2005-01-01

Induction of apoptosis is pivotal for eliminating cells with damaged DNA or deregulated proliferation. We show that tumor suppressor ARF and ATM/ATR kinase pathways cooperate in the induction of apoptosis in response to elevated expression of c-myc, β-catenin or human papilloma virus E7 oncogenes. Overexpression of oncogenes leads to the formation of phosphorylated H2AX foci, induction of Rad51 protein levels and ATM/ATR-dependent phosphorylation of p53. Inhibition of ATM/ATR kinases abolishes both induction of Rad51 and phosphorylation of p53, and remarkably reduces the level of apoptosis induced by co-expression of oncogenes and ARF. However, the induction of apoptosis is downregulated in p53-/- cells and does not depend on activities of ATM/ATR kinases, indicating that efficient induction of apoptosis by oncogene activation depends on coordinated action of ARF and ATM/ATR pathways in the regulation of p53

Possible involvement of MSX-2 homeoprotein in v-ras-induced transformation.

Science.gov (United States)

Takahashi, C; Akiyama, N; Kitayama, H; Takai, S; Noda, M

1997-04-01

A truncated MSX-2 homeoprotein was found to induce flat reversion when expressed in v-Ki-ras-transformed NIH3T3 cells. Although the expression of endogenous MSX-2 gene is low in most of the normal adult tissues examined, it is frequently activated in carcinoma-derived cell lines. Likewise, the gene is inactive in untransformed cells but is transcriptionally activated after transformation by v-Ki-ras oncogene, suggesting that the intact MSX-2 may play a positive, rather than suppressive, role in cell transformation. To test this possibility, we isolated a full-length human MSX-2 cDNA and tested its activities in two cell systems: fibroblast and myoblast. In NIH3T3 fibroblasts, although the gene by itself failed to confer a transformed phenotype, antisense MSX-2 cDNA as well as truncated MSX-2 cDNA interfered with the transforming activities of both v-Ki-ras and v-raf oncogene. In C2C12 myoblasts, MSX-2 was found to suppress MyoD gene expression, as do activated ras oncogenes, under certain culture conditions, and truncated MSX-2 cDNA was found to inhibit the activities of both MSX-2 and ras in this system as well. Our findings not only suggest that the truncated version MSX-2 may act as a dominant suppressor of intact MSX-2 but also raise the possibility that MSX-2 gene may be an important downstream target for the Ras signaling pathways.

Molecular interaction between K-Ras and H-REV107 in the Ras signaling pathway.

Science.gov (United States)

Han, Chang Woo; Jeong, Mi Suk; Jang, Se Bok

2017-09-16

Ras proteins are small GTPases that serve as master moderators of a large number of signaling pathways involved in various cellular processes. Activating mutations in Ras are found in about one-third of cancers. H-REV107, a K-Ras binding protein, plays an important role in determining K-Ras function. H-REV107 is a member of the HREV107 family of class II tumor suppressor genes and a growth inhibitory Ras target gene that suppresses cellular growth, differentiation, and apoptosis. Expression of H-REV107 was strongly reduced in about 50% of human carcinoma cell lines. However, the specific molecular mechanism by which H-REV107 inhibits Ras is still unknown. In the present study, we suggest that H-REV107 forms a strong complex with activating oncogenic mutation Q61H K-Ras from various biochemical binding assays and modeled structures. In addition, the interaction sites between K-Ras and H-REV107 were predicted based on homology modeling. Here, we found that some structure-based mutants of the K-Ras disrupted the complex formation with H-REV107. Finally, a novel molecular mechanism describing K-Ras and H-REV107 binding is suggested and insights into new K-Ras effector target drugs are provided. Copyright © 2017 Elsevier Inc. All rights reserved.

High-Affinity Interaction of the K-Ras4B Hypervariable Region with the Ras Active Site

Science.gov (United States)

Chavan, Tanmay S.; Jang, Hyunbum; Khavrutskii, Lyuba; Abraham, Sherwin J.; Banerjee, Avik; Freed, Benjamin C.; Johannessen, Liv; Tarasov, Sergey G.; Gaponenko, Vadim; Nussinov, Ruth; Tarasova, Nadya I.

2015-01-01

Ras proteins are small GTPases that act as signal transducers between cell surface receptors and several intracellular signaling cascades. They contain highly homologous catalytic domains and flexible C-terminal hypervariable regions (HVRs) that differ across Ras isoforms. KRAS is among the most frequently mutated oncogenes in human tumors. Surprisingly, we found that the C-terminal HVR of K-Ras4B, thought to minimally impact the catalytic domain, directly interacts with the active site of the protein. The interaction is almost 100-fold tighter with the GDP-bound than the GTP-bound protein. HVR binding interferes with Ras-Raf interaction, modulates binding to phospholipids, and slightly slows down nucleotide exchange. The data indicate that contrary to previously suggested models of K-Ras4B signaling, HVR plays essential roles in regulation of signaling. High affinity binding of short peptide analogs of HVR to K-Ras active site suggests that targeting this surface with inhibitory synthetic molecules for the therapy of KRAS-dependent tumors is feasible. PMID:26682817

Exploring environmental causes of altered ras effects: fragmentation plus integration?

Science.gov (United States)

Porta, Miquel; Ayude, Daniel; Alguacil, Juan; Jariod, Manuel

2003-02-01

Mutations in ras genes are the most common abnormality of oncogenes in human cancer and a major example of activation by point mutation. Experimental and epidemiological studies support the notion that Ki-ras activation and expression may be chemically related. We discuss the potential role of several environmental compounds in the induction or promotion of ras mutations in humans, with a focus on exocrine pancreatic cancer, the human tumor with the highest prevalence at diagnosis of Ki-ras mutations. Organochlorine compounds, organic solvents, and coffee compounds may play an indirect role in causing Ki-ras mutations, rather than as direct inducers of the mutations. Although for some organochlorine compounds the induction of point mutations in ras oncogenes cannot be excluded, it seems more likely that the effects of these compounds are mediated through nongenomic or indirectly genotoxic mechanisms of action. Organic solvents also may act via enzymatic induction of ras mutagens or by providing a proliferation advantage to ras-mutated cell clones. In exocrine pancreatic cancer, caffeine, other coffee compounds, or other factors with which coffee drinking is associated could modulate Ki-ras activation by interfering with DNA repair, cell-cycle checkpoints, and apoptosis. Asbestos, cigarette smoking, and some dietary factors also may be involved in the initiation or the promotion of Ki-ras mutations in lung and colon cancers. Further development of the mechanistic scenarios proposed here could contribute to a meaningful integration of biological, clinical, and environmental knowledge on the causes of altered ras effects. Copyright 2003 Wiley-Liss, Inc.

Injection of an antibody against a p21 c-Ha-ras protein inhibits cleavage in axolotl eggs.

OpenAIRE

Baltus, E; Hanocq-Quertier, J; Hanocq, F; Brachet, J

1988-01-01

The presence of a ras protein was demonstrated in cleaving axolotl eggs by selective immunoprecipitation with a polyclonal antibody against a peptide encoded by the c-Ha-ras oncogene, cellular homolog of the v-Ha-ras oncogene of Harvey rat sarcoma virus. Injection of this antibody into axolotl oocytes subjected to progesterone treatment does not prevent meiotic maturation. Injection of the same antibody into a blastomere of axolotl eggs at the 2- or 4-cell stage causes cleavage arrest in the ...

Expression of proto-oncogenes in non-Hodgkin's lymphomas by in situ hybridization with biotinylated DNA probes

International Nuclear Information System (INIS)

Hamatani, Kiyohiro; Yoshida, Kuniko; Abe, Masumi; Shimaoka, Katsutaro; Shiku, Hiroshi; Akiyama, Mitoshi; Kondo, Hisayoshi.

1989-11-01

Expression of six proto-oncogenes (fos, myc, myb, Ki-ras, Ha-ras, and N-ras) in 43 cases of non-Hodgkin's lymphoma was analyzed by means of in situ hybridization. Biotinylated DNA probes of the six oncogenes and those of the immunoglobulin H-chain (IgH) gene and the T cell receptor β-chain (TCRβ) gene were used. The results of in situ hybridization performed under blind conditions by IgH and TCRβ gene probes were compatible with those of typing by cell surface markers. The nuclear protein-related proto-oncogenes, fos myc, and myb, were expressed in about 70 % - 80 % of all cases regardless of phenotypes, histology or histologic grade. On the contrary, genes of the ras family were expressed in fewer cases except for the Ki-ras gene which was more frequently expressed by cases of the T cell immunophenotype with a high malignancy grade. The results of dot hybridization with RNA extracted from some cases were compatible with those of in situ hybridization, further demonstrating the specificity of in situ hybridization. (author)

Guanosine triphosphatase activating protein (GAP) interacts with the p21 ras effector binding domain

DEFF Research Database (Denmark)

Adari, H; Lowy, D R; Willumsen, B M

1988-01-01

A cytoplasmic protein that greatly enhances the guanosine triphosphatase (GTPase) activity of N-ras protein but does not affect the activity of oncogenic ras mutants has been recently described. This protein (GAP) is shown here to be ubiquitous in higher eukaryotes and to interact with H-ras as w...

PAK1 is a breast cancer oncogene that coordinately activates MAPK and MET signaling.

Science.gov (United States)

Shrestha, Y; Schafer, E J; Boehm, J S; Thomas, S R; He, F; Du, J; Wang, S; Barretina, J; Weir, B A; Zhao, J J; Polyak, K; Golub, T R; Beroukhim, R; Hahn, W C

2012-07-19

Activating mutations in the RAS family or BRAF frequently occur in many types of human cancers but are rarely detected in breast tumors. However, activation of the RAS-RAF-MEK-ERK MAPK pathway is commonly observed in human breast cancers, suggesting that other genetic alterations lead to activation of this signaling pathway. To identify breast cancer oncogenes that activate the MAPK pathway, we screened a library of human kinases for their ability to induce anchorage-independent growth in a derivative of immortalized human mammary epithelial cells (HMLE). We identified p21-activated kinase 1 (PAK1) as a kinase that permitted HMLE cells to form anchorage-independent colonies. PAK1 is amplified in several human cancer types, including 30--33% of breast tumor samples and cancer cell lines. The kinase activity of PAK1 is necessary for PAK1-induced transformation. Moreover, we show that PAK1 simultaneously activates MAPK and MET signaling; the latter via inhibition of merlin. Disruption of these activities inhibits PAK1-driven anchorage-independent growth. These observations establish PAK1 amplification as an alternative mechanism for MAPK activation in human breast cancer and credential PAK1 as a breast cancer oncogene that coordinately regulates multiple signaling pathways, the cooperation of which leads to malignant transformation.

Oncogenes and radiosensitivity: in vitro studies. Potential impact in radiotherapy

International Nuclear Information System (INIS)

Alapetite, C.; Moustacchi, E.; Cosset, J.M.

1992-01-01

It is of interest to address the question of whether or not activated oncogenes can influence tumorigenic cell response to radiations. Malignant transformation through transfection of oncogenes offers a possibility for in vitro comparison of transformed cells and parental cells. Murin cellular system analysis suggests an acquisition of radioresistance through some oncogenes transfection. In human cells, only a limited number of oncogenes (ras and myc) has been studied so far. To date, no crucial influence could be demonstrated. The extension of the analysis to other oncogenes and suppressor genes could potentially be helpful for the choice and the modalities of cancer treatment

c-Ha-ras BamHI RFLP in human urothelial tumors and point mutations in hot codons

International Nuclear Information System (INIS)

Weismanova, E; Skovraga, M.; Kaluz, S.

1993-01-01

High-molecular weights DNAs from 30 bladder and renal cell carcinomas (RCC) were isolated and the c-Ha-ras the c-Ha-ras gene BamHI RFLP was examined. Amplification of c-Ha-ras with normal localization with regard to the size of alleles was found only in the case. One of the normally localized c-Ha-ras allele termed RCC c-H-ras of a length of about 6.6 kbp was cloned and an oncogene-activating point mutation was identified using two restriction enzymes. After comparison of CfrI and Cfr10I cleavage maps of RCC c-Ha-ras to complete nucleotide sequences of EJ/T24 c-Ha-ras oncogene and its normal counterpart, a point mutation was identified within codon 11 or 12. The use of CfrI and Cfr10I is of value for clinical practice in identification of point mutations in c-Ha-ras PCR product in neoplasia accompanied by somatic mutation of c-Ha-ras. The correlation among c-Ha-ras allele, amplification/loss, presence of point mutation and progression of neoplasia is discussed. (author)

Characterization of ERAS, a putative novel human oncogene, in skin and breast

Energy Technology Data Exchange (ETDEWEB)

Peña Avalos, B.L. de la

2014-07-01

Most human tumors have mutations in genes of the RAS small GTPase protein family. RAS works as a molecular switch for signaling pathways that modulate many aspects of cell behavior, including proliferation, differentiation, motility and death. Oncogenic mutations in RAS prevent GTP hydrolysis, locking RAS in a permanently active state, being the most common mutations in HRAS, KRAS and NRAS. The human RAS family consists of at least 36 different genes, many of which have been scarcely studied. One of these relatively unknown genes is ERAS (ES cell-expressed RAS), which is a constitutively active RAS protein, localized in chromosome X and expressed only in embryonic cells, being undetectable in adult tissues. New high throughput technologies have made it possible to screen complete cancer genomes for identification of mutations associated to cancer. Using the Sleeping Beauty (SB) transposon system, ERAS was identified as a putative novel oncogene in non-melanoma skin and breast cancers. The major aim of this project is to determine the general characteristics of ERAS as a putative novel human oncogene in skin and breast cells. Forced expression of ERAS results in drastic changes in cell shape, proliferation and motility. When ERAS is overexpressed in skin and breast human cells it is mainly localized in the cytoplasmic membrane. ERAS activates the phosphatidylinositol-3-OH kinase (PI3K) pathway but not the mitogen-activated protein kinase (MAPK) pathway. ERAS-expressing cells suffer spontaneous morphologic and phenotypic EMT-like changes, including cytoskeleton reorganization, vimentin and N-cadherin up-regulation and down-regulation of E-cadherin, which can be associated with increased malignancy, and invasive and metastatic potential. Our results suggest that inappropriate expression of ERAS lead to transformation of human cells. (Author)

A Novel Ras Effector Pathway Found to Play Significant Role in Tumor Suppression | Poster

Science.gov (United States)

By Nancy Parrish, Staff Writer; photo by Richard Frederickson, Staff Photographer Normal cells have mechanisms to prevent the development of cancer. Among these is a type of tumor suppressor mechanism known as oncogene-induced senescence, or OIS, which halts the uncontrolled growth of cells caused by mutations in oncogenes. The oncogene Ras plays a crucial role in inducing OIS

Cellular oncogene expression following exposure of mice to γ-rays

International Nuclear Information System (INIS)

Anderson, A.; Woloschak, G.E.

1991-01-01

We examined the effects of total body exposure of BCF1 mice to γ-rays (300 cGy) in modulating expression of cellular oncogenes in both gut and liver tissues. We selected specific cellular oncogenes (c-fos, c-myc, c-src, and c-H-ras), based on their normal expression in liver and gut tissues from untreated mice. As early as 5 min. following whole body exposure of BCF1 mice to γ-rays we detected induction of mRNA specific for c-src and c-H-ras in both liver and gut tissues. c-fos RNA was slightly decreased in accumulation in gut but was unaffected in liver tissue from irradiated mice relative to untreated controls. c-myc mRNA accumulation was unaffected in all tissues examined. These experiments document that modulation of cellular oncogene expression can occur as an early event in tissues following irradiation and suggest that this modulation may play a role in radiation-induced carcinogenesis

Genetic analysis of Ras genes in epidermal development and tumorigenesis

Science.gov (United States)

Drosten, Matthias; Lechuga, Carmen G; Barbacid, Mariano

2013-01-01

Proliferation and differentiation of epidermal keratinocytes are tightly controlled to ensure proper development and homeostasis of the epidermis. The Ras family of small GTPases has emerged as a central node in the coordination of cell proliferation in the epidermis. Recent genetic evidence from mouse models has revealed that the intensity of Ras signaling modulates the proliferative capacity of epidermal keratinocytes. Interfering with Ras signaling either by combined elimination of the 3 Ras genes from the basal layer of the epidermis or by overexpression of dominant-negative Ras isoforms caused epidermal thinning due to hypoproliferation of keratinocytes. In contrast, overexpression of oncogenic Ras mutants in different epidermal cell layers led to hyperproliferative phenotypes including the development of papillomas and squamous cell carcinomas. Here, we discuss the value of loss- and gain-of-function studies in mouse models to assess the role of Ras signaling in the control of epidermal proliferation. PMID:24150175

ΔNp63α is an oncogene that induces Lsh expression and promotes stem-like proliferation

Science.gov (United States)

Keyes, William M.; Pecoraro, Matteo; Aranda, Victoria; Vernersson-Lindahl, Emma; Li, Wangzhi; Vogel, Hannes; Guo, Xuecui; Garcia, Elvin L.; Michurina, Tatyana V.; Enikolopov, Grigori; Muthuswamy, Senthil K.; Mills, Alea A.

2014-01-01

SUMMARY The p53 homolog p63 is essential for development, yet its role in cancer is not clear. We discovered that p63 deficiency evokes the tumor suppressive mechanism of cellular senescence, causing a striking absence of stratified epithelia such as the skin. Here we identify the predominant p63 isoform, ΔNp63α, as a protein that bypasses oncogene induced senescence to drive tumorigenesis in vivo. Interestingly, bypass of senescence promotes stem-like proliferation and maintains survival of the keratin 15-positive stem cell population. Furthermore, we identify the chromatin remodeling protein Lsh as a new target of ΔNp63α that is an essential mediator of senescence bypass. These findings indicate that ΔNp63α is an oncogene that cooperates with Ras to promote tumor-initiating stem-like proliferation, and suggest that Lsh-mediated chromatin remodeling events are critical to this process. PMID:21295273

Mutant p53 - heat shock response oncogenic cooperation: a new mechanism of cancer cell survival

Directory of Open Access Journals (Sweden)

Evguenia eAlexandrova

2015-04-01

Full Text Available The main tumor suppressor function of p53 as a ‘guardian of the genome’ is to respond to cellular stress by transcriptional activation of apoptosis, growth arrest or senescence in damaged cells. Not surprisingly, mutations in the p53 gene are the most frequent genetic alteration in human cancers. Importantly, mutant p53 (mutp53 proteins not only lose their wild-type tumor suppressor activity, but also can actively promote tumor development. Two main mechanisms accounting for mutp53 proto-oncogenic activity are inhibition of the wild-type p53 in a dominant-negative fashion and gain of additional oncogenic activities known as gain-of-function (GOF. Here we discuss a novel mechanism of mutp53 GOF, which relies on its oncogenic cooperation with the heat shock machinery. This coordinated adaptive mechanism renders cancer cells more resistant to proteotoxic stress and provides both, a strong survival advantage to cancer cells and a promising means for therapeutic intervention.

PI3K: A Crucial Piece in the RAS Signaling Puzzle.

Science.gov (United States)

Krygowska, Agata Adelajda; Castellano, Esther

2018-06-01

RAS proteins are key signaling switches essential for control of proliferation, differentiation, and survival of eukaryotic cells. RAS proteins are mutated in 30% of human cancers. In addition, mutations in upstream or downstream signaling components also contribute to oncogenic activation of the pathway. RAS proteins exert their functions through activation of several signaling pathways and dissecting the contributions of these effectors in normal cells and in cancer is an ongoing challenge. In this review, we summarize our current knowledge about how RAS regulates type I phosphatidylinositol 3-kinase (PI3K), one of the main RAS effectors. RAS signaling through PI3K is necessary for normal lymphatic vasculature development and for RAS-induced transformation in vitro and in vivo, especially in lung cancer, where it is essential for tumor initiation and necessary for tumor maintenance. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

Limited role of murine ATM in oncogene-induced senescence and p53-dependent tumor suppression.

Directory of Open Access Journals (Sweden)

Alejo Efeyan

Full Text Available Recent studies in human fibroblasts have provided a new general paradigm of tumor suppression according to which oncogenic signaling produces DNA damage and this, in turn, results in ATM/p53-dependent cellular senescence. Here, we have tested this model in a variety of murine experimental systems. Overexpression of oncogenic Ras in murine fibroblasts efficiently induced senescence but this occurred in the absence of detectable DNA damage signaling, thus suggesting a fundamental difference between human and murine cells. Moreover, lung adenomas initiated by endogenous levels of oncogenic K-Ras presented abundant senescent cells, but undetectable DNA damage signaling. Accordingly, K-Ras-driven adenomas were also senescent in Atm-null mice, and the tumorigenic progression of these lesions was only modestly accelerated by Atm-deficiency. Finally, we have examined chemically-induced fibrosarcomas, which possess a persistently activated DNA damage response and are highly sensitive to the activity of p53. We found that the absence of Atm favored genomic instability in the resulting tumors, but did not affect the persistent DNA damage response and did not impair p53-dependent tumor suppression. All together, we conclude that oncogene-induced senescence in mice may occur in the absence of a detectable DNA damage response. Regarding murine Atm, our data suggest that it plays a minor role in oncogene-induced senescence or in p53-dependent tumor suppression, being its tumor suppressive activity probably limited to the maintenance of genomic stability.

The oncogenic action of ionizing radiation on rat skin

International Nuclear Information System (INIS)

Burns, F.J.

1991-01-01

Progress has occurred in several areas corresponding to the specific aims of the proposal: (1) Progression and multiple events in radiation carcinogenesis of rat skin as a function of LET; (2) cell cycle kinetics of irradiated rat epidermis as determined by double labeling and double emulsion autoradiography; (3) oncogene activation detected by in situ hybridization in radiation-induced rat skin tumors; (4) amplification of the c-myc oncogene in radiation-induced rat skin tumors as a function of LET; and (5) transformation of rat skin keratinocytes by ionizing radiation in combination with c-Ki-ras and c-myc oncogenes. 111 refs., 13 figs., 12 tabs

The radiosensitivity of human keratinocytes: influence of activated c-H-ras oncogene expression and tumorigenicity

International Nuclear Information System (INIS)

Mendonca, M.S.; Redpath, J.L.; Stanbridge, E.J.

1991-01-01

The authors investigated γ-ray sensitivity of several activated c-H-ras (EJ) containing clones established after transfection of the spontaneously immortalized non-tumorigenic human keratinocyte cell line HaCaT. The clones were grouped according to tumorigenic potential after subcutaneous injection into nude mice, and fell into three classes: Class I clones A-4 and I-6 are non-tumorigenic and express very low levels of c-H-ras mRNA and no mutated ras protein (p 21 ); Class II clones I-5 and I-7 grow to large (benign) epidermal cysts, express intermediate to high c-H-ras mRNA and variable levels of mutated ras p 21 protein with clone I-5 expressing little and clone I-7 expressing high levels of p 21 ; Class III clones II-3 and II-4 grow to solid squamous cell carcinomas, express high c-H-ras mRNA and high level of mutated p 21 ras protein similar to clone I-7. Comparison of single-hit multitarget or linear-quadratic survival curve parameters, and survival at 2Gy (S 2 ) indicate no general correlation with either activated c-H-ras expression level or tumorigenic potential, and increased radioresistance. (author)

The role of autophagy in cytotoxicity induced by new oncogenic B-Raf inhibitor UI-152 in v-Ha-ras transformed fibroblasts

International Nuclear Information System (INIS)

Ahn, Jun-Ho; Ahn, Soon Kil; Lee, Michael

2012-01-01

Highlights: ► We recently discovered a potent and selective B-Raf inhibitor, UI-152. ► UI-152 displayed a selective cytotoxicity toward v-Ha-ras transformed cells. ► UI-152-induced growth inhibition was largely meditated by autophagy. ► UI-152 induced paradoxical activation of Raf-1. -- Abstract: In human cancers, B-Raf is the most frequently mutated protein kinase in the MAPK signaling cascade, making it an important therapeutic target. We recently discovered a potent and selective B-Raf inhibitor, UI-152, by using a structure-based drug design strategy. In this study, we examined whether B-Raf inhibition by UI-152 may be an effective therapeutic strategy for eliminating cancer cells transformed with v-Ha-ras (Ras-NIH 3T3). UI-152 displayed selective cytotoxicity toward Ras-NIH 3T3 cells while having little to no effect on non-transformed NIH 3T3 cells. We found that treatment with UI-152 markedly increased autophagy and, to a lesser extent, apoptosis. However, inhibition of autophagy by addition of 3-MA failed to reverse the cytotoxic effects of UI-152 on Ras-NIH 3T3 cells, demonstrating that apoptosis and autophagy can act as cooperative partners to induce growth inhibition in Ras-NIH 3T3 cells treated with UI-152. Most interestingly, cell responses to UI-152 appear to be paradoxical. Here, we showed that although UI-152 inhibited ERK, it induced B-Raf binding to Raf-1 as well as Raf-1 activation. This paradoxical activation of Raf-1 by UI-152 is likely to be coupled with the inhibition of the mTOR pathway, an intracellular signaling pathway involved in autophagy. We also showed for the first time that, in multi-drug resistant cells, the combination of UI-152 with verapamil significantly decreased cell proliferation and increased autophagy. Thus, our findings suggest that the inhibition of autophagy, in combination with UI-152, offers a more effective therapeutic strategy for v-Ha-ras-transformed cells harboring wild-type B-Raf.

Oncogene activation and surface markers in mouse lymphomas induced by radiation and nitrosomethylurea

Energy Technology Data Exchange (ETDEWEB)

Guerrero, I.; Villasante, A.; Diamond, L.; Berman, J.W.; Newcomb, E.W.; Steinberg, J.J.; Lake, R.; Pellicer, A.

1986-01-01

Thymic lymphomas have been induced by ..gamma..-radiation and treatment with the chemical nitrosomethylurea in different mice strains. As indicated by the NIH 3T3 focus forming assay, a significant percentage of the tumors contain activated oncogenes of the ras family (K or N). Cloning and sequencing has enabled us to identify single base mutations as the only significant alteration present in the activated oncogenes. These alterations result in the substitution of amino-acid 12 or 61 of the p21 product of the ras genes. With the use of synthetic oligonucleotides it has been found that the tumors do not all contain the same mutation and in one case so far the normal allele is absent.

Decomposing Oncogenic Transcriptional Signatures to Generate Maps of Divergent Cellular States.

Science.gov (United States)

Kim, Jong Wook; Abudayyeh, Omar O; Yeerna, Huwate; Yeang, Chen-Hsiang; Stewart, Michelle; Jenkins, Russell W; Kitajima, Shunsuke; Konieczkowski, David J; Medetgul-Ernar, Kate; Cavazos, Taylor; Mah, Clarence; Ting, Stephanie; Van Allen, Eliezer M; Cohen, Ofir; Mcdermott, John; Damato, Emily; Aguirre, Andrew J; Liang, Jonathan; Liberzon, Arthur; Alexe, Gabriella; Doench, John; Ghandi, Mahmoud; Vazquez, Francisca; Weir, Barbara A; Tsherniak, Aviad; Subramanian, Aravind; Meneses-Cime, Karina; Park, Jason; Clemons, Paul; Garraway, Levi A; Thomas, David; Boehm, Jesse S; Barbie, David A; Hahn, William C; Mesirov, Jill P; Tamayo, Pablo

2017-08-23

The systematic sequencing of the cancer genome has led to the identification of numerous genetic alterations in cancer. However, a deeper understanding of the functional consequences of these alterations is necessary to guide appropriate therapeutic strategies. Here, we describe Onco-GPS (OncoGenic Positioning System), a data-driven analysis framework to organize individual tumor samples with shared oncogenic alterations onto a reference map defined by their underlying cellular states. We applied the methodology to the RAS pathway and identified nine distinct components that reflect transcriptional activities downstream of RAS and defined several functional states associated with patterns of transcriptional component activation that associates with genomic hallmarks and response to genetic and pharmacological perturbations. These results show that the Onco-GPS is an effective approach to explore the complex landscape of oncogenic cellular states across cancers, and an analytic framework to summarize knowledge, establish relationships, and generate more effective disease models for research or as part of individualized precision medicine paradigms. Copyright © 2017 Elsevier Inc. All rights reserved.

Differential p53 engagement in response to oxidative and oncogenic stresses in Fanconi anemia mice

Science.gov (United States)

Rani, Reena; Li, Jie; Pang, Qishen

2008-01-01

Members of the Fanconi anemia (FA) protein family are involved in repair of genetic damage caused by DNA cross-linkers. It is not clear whether the FA proteins function in oxidative DNA damage and oncogenic stress response. Here we report that deficiency in the Fanca gene in mice elicits a p53-dependent growth arrest and DNA damage response to oxidative DNA damage and oncogenic stress. Using a Fanca-/- Trp53-/- double knockout model and a functionally switchable p53 retrovirus, we define the kinetics, dependence, and persistence of p53-mediated response to oxidative and oncogenic stresses in Fanca-/- cells. Notably, oxidative stress induces persistent p53 response in Fanca-/- cells, likely due to accumulation of unrepaired DNA damage. On the other hand, whereas WT cells exhibit prolonged response to oncogene activation, the p53-activating signals induced by oncogenic ras are short-lived in Fanca-/- cells, suggesting that Fanca may be required for the cell to engage p53 during constitutive ras activation. We propose that the FA proteins protect cells from stress-induced proliferative arrest and tumor evolution by acting as a modulator of the signaling pathways that link FA to p53. PMID:19047147

Differential p53 engagement in response to oxidative and oncogenic stresses in Fanconi anemia mice.

Science.gov (United States)

Rani, Reena; Li, Jie; Pang, Qishen

2008-12-01

Members of the Fanconi anemia (FA) protein family are involved in repair of genetic damage caused by DNA cross-linkers. It is not clear whether the FA proteins function in oxidative DNA damage and oncogenic stress response. Here, we report that deficiency in the Fanca gene in mice elicits a p53-dependent growth arrest and DNA damage response to oxidative DNA damage and oncogenic stress. Using a Fanca-/-Trp53-/- double knockout model and a functionally switchable p53 retrovirus, we define the kinetics, dependence, and persistence of p53-mediated response to oxidative and oncogenic stresses in Fanca-/- cells. Notably, oxidative stress induces persistent p53 response in Fanca-/- cells, likely due to accumulation of unrepaired DNA damage. On the other hand, whereas wild-type cells exhibit prolonged response to oncogene activation, the p53-activating signals induced by oncogenic ras are short-lived in Fanca-/- cells, suggesting that Fanca may be required for the cell to engage p53 during constitutive ras activation. We propose that the FA proteins protect cells from stress-induced proliferative arrest and tumor evolution by acting as a modulator of the signaling pathways that link FA to p53.

Oncogene expression in primary lung tumors in dogs that inhaled 239PuO2

International Nuclear Information System (INIS)

Kelly, G.; Kerkof, P.R.; Haley, P.J.

1988-01-01

Ten radiation-induced and three spontaneous lung tumors were analyzed for aberrant expression of known oncogenes. In 12 of 13 tumors tested, sequences hybridizing to the c-myc oncogene were expressed at levels 1.5 times higher than sequences hybridizing to β-actin. This level of oncogene expression was also observed in 9 of 13 tumors for 1 or more members of the ras family of oncogenes. Seven of thirteen tumors examined express sequences that hybridize with clones of v-ros or c-met. The ros and met clones both code for oncogenes whose normal homologues are transmembrane proteins related to the insulin receptor. (author)

Novel approach to abuse the hyperactive K-Ras pathway for adenoviral gene therapy of colorectal cancer

International Nuclear Information System (INIS)

Naumov, Inna; Kazanov, Dina; Lisiansky, Victoria; Starr, Alex; Aroch, Ilan; Shapira, Shiran; Kraus, Sarah; Arber, Nadir

2012-01-01

Background: Functional activation of oncogenic K-Ras signaling pathway plays an important role in the early events of colorectal carcinogenesis (CRC). K-Ras proto-oncogene is involved in 35–40% of CRC cases. Mutations in the Ras gene trigger the transduction of proliferative and anti-apoptotic signals, even in the absence of extra cellular stimuli. The objective of the current study was to use a gene-targeting approach to kill human CRC cells selectively harboring mutated K-Ras. Results: A recombinant adenovirus that carries a lethal gene, PUMA, under the control of a Ras responsive promoter (Ad-Py4-SV40-PUMA) was used selectively to target CRC cells (HCT116, SW480, DLD1 and RIE-Ras) that possess a hyperactive Ras pathway while using HT29 and RIE cells as a control that harbors wild type Ras and exhibit very low Ras activity. Control vector, without the Ras responsive promoter elements was used to assess the specificity of our “gene therapy” approach. Both adenoviral vectors were assed in vitro and in xenograft model in vivo. Ad-Py4-SV40-PUMA showed high potency to induce ∼ 50% apoptosis in vitro, to abolish completely tumor formation by infecting cells with the Ad-Py4-SV40-PUMA prior xenografting them in nude mice and high ability to suppress by ∼ 35% tumor progression in vivo in already established tumors. Conclusions: Selective targeting of CRC cells with the activated Ras pathway may be a novel and effective therapy in CRC. The high potency of this adenoviral vector may help to overcome an undetectable micro metastasis that is the major hurdle in challenging with CRC.

Novel approach to abuse the hyperactive K-Ras pathway for adenoviral gene therapy of colorectal cancer

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Naumov, Inna [Integrated Cancer Prevention Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel); Kazanov, Dina [Integrated Cancer Prevention Center, Tel Aviv (Israel); Lisiansky, Victoria [Integrated Cancer Prevention Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel); Starr, Alex [Lung and Allergy Institute, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel); Aroch, Ilan; Shapira, Shiran; Kraus, Sarah [Integrated Cancer Prevention Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel); Arber, Nadir, E-mail: narber@post.tau.ac.il [Integrated Cancer Prevention Center, Tel Aviv (Israel); Department of Gastroenterology, Tel Aviv Sourasky Medical Center, Tel Aviv (Israel); Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv (Israel)

2012-01-15

Background: Functional activation of oncogenic K-Ras signaling pathway plays an important role in the early events of colorectal carcinogenesis (CRC). K-Ras proto-oncogene is involved in 35-40% of CRC cases. Mutations in the Ras gene trigger the transduction of proliferative and anti-apoptotic signals, even in the absence of extra cellular stimuli. The objective of the current study was to use a gene-targeting approach to kill human CRC cells selectively harboring mutated K-Ras. Results: A recombinant adenovirus that carries a lethal gene, PUMA, under the control of a Ras responsive promoter (Ad-Py4-SV40-PUMA) was used selectively to target CRC cells (HCT116, SW480, DLD1 and RIE-Ras) that possess a hyperactive Ras pathway while using HT29 and RIE cells as a control that harbors wild type Ras and exhibit very low Ras activity. Control vector, without the Ras responsive promoter elements was used to assess the specificity of our 'gene therapy' approach. Both adenoviral vectors were assed in vitro and in xenograft model in vivo. Ad-Py4-SV40-PUMA showed high potency to induce {approx} 50% apoptosis in vitro, to abolish completely tumor formation by infecting cells with the Ad-Py4-SV40-PUMA prior xenografting them in nude mice and high ability to suppress by {approx} 35% tumor progression in vivo in already established tumors. Conclusions: Selective targeting of CRC cells with the activated Ras pathway may be a novel and effective therapy in CRC. The high potency of this adenoviral vector may help to overcome an undetectable micro metastasis that is the major hurdle in challenging with CRC.

The role of hypoxia inducible factor-1 alpha in bypassing oncogene-induced senescence.

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Mehtap Kilic Eren

Full Text Available Oncogene induced senescence (OIS is a sustained anti-proliferative response acutely induced in primary cells via activation of mitogenic oncogenes such as Ras/BRAF. This mechanism acts as an initial barrier preventing normal cells transformation into malignant cell. Besides oncogenic activation and DNA damage response (DDR, senescence is modulated by a plethora of other factors, and one of the most important one is oxygen tension of the tissue. The aim of this study was to determine the impact of hypoxia on RasV12-induced senescence in human diploid fibroblasts (HDFs. We showed here that hypoxia prevents execution of oncogene induced senescence (OIS, through a strong down-regulation of senescence hallmarks, such as SA- β-galactosidase, H3K9me3, HP1γ, p53, p21CIP1 and p16INK4a in association with induction of hypoxia inducible factor-1α (HIF-1α. In addition, hypoxia also decreased marks of H-RasV12-induced DDR in both cell lines through down-regulation of ATM/ATR, Chk1 and Chk2 phosphorylation as well as decreased γ-H2AX positivity. Utilizing shRNA system targeting HIF-1α we show that HIF-1α is directly involved in down regulation of p53 and its target p21CIP1 but not p16INK4a. In line with this finding we found that knock down of HIF-1α leads to a strong induction of apoptotic response, but not restoration of senescence in Ras expressing HDFs in hypoxia. This indicates that HIF-1α is an important player in early steps of tumorigenesis, leading to suppression of senescence through its negative regulation of p53 and p21CIP1. In our work we describe a mechanism through which hypoxia and specifically HIF-1α preclude cells from maintaining senescence-driven anti proliferative response. These findings indicate the possible mechanism through which hypoxic environment helps premalignant cells to evade impingement of cellular failsafe pathways.

The inhibitory NKR-P1B:Clr-b recognition axis facilitates detection of oncogenic transformation and cancer immunosurveillance

DEFF Research Database (Denmark)

Tanaka, M; Fine, Jason; Kirkham, Christina

2018-01-01

Natural killer (NK) cells express receptors specific for MHC class I (MHC-I) molecules involved in "missing-self" recognition of cancer and virus-infected cells. Here we elucidate the role of MHC-I-independent NKR-P1B:Clr-b interactions in the detection of oncogenic transformation by NK cells. Ras......-b protein, in turn promoting missing-self recognition via the NKR-P1B inhibitory receptor. Both Ras- and c-Myc-mediated Clr-b loss selectively augmented cytotoxicity of oncogene-transformed leukemia cells by NKR-P1B+ NK cells in vitro and enhanced rejection by WT mice in vivo. Interestingly, genetic...

THE MYC FAMILY OF ONCOGENES AND THEIR PRESENCE AND IMPORTANCE IN SMALL-CELL LUNG-CARCINOMA AND OTHER TUMOR TYPES

NARCIS (Netherlands)

DEVRIES, EGE; MULDER, NH

1993-01-01

The myc family of cellular oncogenes, c - myr, N - myc, encodes three highly related, cell cycle specific, nuclear phosphoproteins. All are able to transform primary rat embryo fibroblasts when cotransfected with the c - ras oncogene. Myc family genes am differentially expressed with respect to

Oncogene expression in primary lung tumors in dogs that inhaled {sup 239}PuO{sub 2}

Energy Technology Data Exchange (ETDEWEB)

Kelly, G; Kerkof, P R; Haley, P J

1988-12-01

Ten radiation-induced and three spontaneous lung tumors were analyzed for aberrant expression of known oncogenes. In 12 of 13 tumors tested, sequences hybridizing to the c-myc oncogene were expressed at levels 1.5 times higher than sequences hybridizing to {beta}-actin. This level of oncogene expression was also observed in 9 of 13 tumors for 1 or more members of the ras family of oncogenes. Seven of thirteen tumors examined express sequences that hybridize with clones of v-ros or c-met. The ros and met clones both code for oncogenes whose normal homologues are transmembrane proteins related to the insulin receptor. (author)

P120-GAP associated with syndecan-2 to function as an active switch signal for Src upon transformation with oncogenic ras

International Nuclear Information System (INIS)

Huang, J.-W.; Chen, C.-L.; Chuang, N.-N.

2005-01-01

BALB/3T3 cells transfected with plasmids pcDNA3.1-[S-ras(Q 61 K)] of shrimp Penaeus japonicus were applied to reveal a complex of p120-GAP/syndecan-2 being highly expressed upon transformation. Of interest, most of the p120-GAP/syndecan-2 complex was localized at caveolae, a membrane microdomain enriched with caveolin-1. To confirm the molecular interaction between syndecan-2 and p120-GAP, we further purified p120-GAP protein from mouse brains by using an affinity column of HiTrap-RACK1 and expressed mouse RACK1-encoded fusion protein and mouse syndecan-2-encoded fusion protein in bacteria. We report molecular affinities exist between p120-GAP and RACK1, syndecan-2 and RACK1 as well as p120-GAP and syndecan-2. The selective affinity between p120-GAP and syndecan-2 was found to be sufficient to detach RACK1. The p120-GAP/syndecan-2 complex was demonstrated to keep Src tyrosine kinase in an activated form. On the other hand, the syndecan-2/RACK1 complex was found to have Src in an inactivated form. These data indicate that the p120-GAP/syndecan-2 complex at caveolae could provide a docking site for Src to transmit tyrosine signaling, implying that syndecan-2/p120-GAP functions as a tumor promoter upon transformation with oncogenic ras of shrimp P. japonicus

Multistep carcinogenesis of normal human fibroblasts. Human fibroblasts immortalized by repeated treatment with Co-60 gamma rays were transformed into tumorigenic cells with Ha-ras oncogenes.

Science.gov (United States)

Namba, M; Nishitani, K; Fukushima, F; Kimoto, T

1988-01-01

Two normal mortal human fibroblast cell strains were transformed into immortal cell lines, SUSM-1 and KMST-6, by treatment with 4-nitroquinoline 1-oxide (4NQO) and Co-60 gamma rays, respectively. These immortalized cell lines showed morphological changes of cells and remarkable chromosome aberrations, but neither of them grew in soft agar or formed tumors in nude mice. The immortal cell line, KMST-6, was then converted into neoplastic cells by treatment with Harvey murine sarcoma virus (Ha-MSV) or the c-Ha-ras oncogene derived from a human lung carcinoma. These neoplastically transformed cells acquired anchorage-independent growth potential and developed tumors when transplanted into nude mice. All the tumors grew progressively without regression until the animals died of tumors. In addition, the tumors were transplantable into other nude mice. Normal human fibroblasts, on the other hand, were not transformed into either immortal or tumorigenic cells by treatment with Ha-MSV or c-Ha-ras alone. Our present data indicate that (1) the chemical carcinogen, 4NQO, or gamma rays worked as an initiator of carcinogenesis in normal human cells, giving rise to immortality, and (2) the ras gene played a role in the progression of the immortally transformed cells to more malignant cells showing anchorage-independent growth and tumorigenicity. In other words, the immortalization process of human cells seems to be a pivotal or rate-limiting step in the carcinogenesis of human cells.

Oncogene Mimicry as a Mechanism of Primary Resistance to BRAF Inhibitors

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Martin L. Sos

2014-08-01

Full Text Available Despite the development of potent RAF/mitogen-activated protein kinase (MAPK pathway inhibitors, only a fraction of BRAF-mutant patients benefit from treatment with these drugs. Using a combined chemogenomics and chemoproteomics approach, we identify drug-induced RAS-RAF-MEK complex formation in a subset of BRAF-mutant cancer cells characterized by primary resistance to vemurafenib. In these cells, autocrine interleukin-6 (IL-6 secretion may contribute to the primary resistance phenotype via induction of JAK/STAT3 and MAPK signaling. In a subset of cell lines, combined IL-6/MAPK inhibition is able to overcome primary resistance to BRAF-targeted therapy. Overall, we show that the signaling plasticity exerted by primary resistant BRAF-mutant cells is achieved by their ability to mimic signaling features of oncogenic RAS, a strategy that we term “oncogene mimicry.” This model may guide future strategies for overcoming primary resistance observed in these tumors.

Approach for targeting Ras with small molecules that activate SOS-mediated nucleotide exchange.

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Burns, Michael C; Sun, Qi; Daniels, R Nathan; Camper, DeMarco; Kennedy, J Phillip; Phan, Jason; Olejniczak, Edward T; Lee, Taekyu; Waterson, Alex G; Rossanese, Olivia W; Fesik, Stephen W

2014-03-04

Aberrant activation of the small GTPase Ras by oncogenic mutation or constitutively active upstream receptor tyrosine kinases results in the deregulation of cellular signals governing growth and survival in ∼30% of all human cancers. However, the discovery of potent inhibitors of Ras has been difficult to achieve. Here, we report the identification of small molecules that bind to a unique pocket on the Ras:Son of Sevenless (SOS):Ras complex, increase the rate of SOS-catalyzed nucleotide exchange in vitro, and modulate Ras signaling pathways in cells. X-ray crystallography of Ras:SOS:Ras in complex with these molecules reveals that the compounds bind in a hydrophobic pocket in the CDC25 domain of SOS adjacent to the Switch II region of Ras. The structure-activity relationships exhibited by these compounds can be rationalized on the basis of multiple X-ray cocrystal structures. Mutational analyses confirmed the functional relevance of this binding site and showed it to be essential for compound activity. These molecules increase Ras-GTP levels and disrupt MAPK and PI3K signaling in cells at low micromolar concentrations. These small molecules represent tools to study the acute activation of Ras and highlight a pocket on SOS that may be exploited to modulate Ras signaling.

PAK1 is a breast cancer oncogene that coordinately activates MAPK and MET signaling

OpenAIRE

Shrestha, Yashaswi; Schafer, Eric J.; Boehm, Jesse S.; Thomas, Sapana R.; He, Frank; Du, Jinyan; Wang, Shumei; Barretina, Jordi; Weir, Barbara A.; Zhao, Jean J.; Polyak, Kornelia; Golub, Todd R.; Beroukhim, Rameen; Hahn, William C.

2011-01-01

Activating mutations in the RAS family or BRAF frequently occur in many types of human cancers but are rarely detected in breast tumors. However, activation of the RAS-RAF-MEK-ERK Mitogen-Activated Protein Kinase (MAPK) pathway is commonly observed in human breast cancers, suggesting that other genetic alterations lead to activation of this signaling pathway. To identify breast cancer oncogenes that activate the MAPK pathway, we screened a library of human kinases for their ability to induce ...

The Bisphenol A analogue Bisphenol S binds to K-Ras4B--implications for 'BPA-free' plastics.

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Schöpel, Miriam; Herrmann, Christian; Scherkenbeck, Jürgen; Stoll, Raphael

2016-02-01

K-Ras4B is a small GTPase that belongs to the Ras superfamily of guanine nucleotide-binding proteins. GTPases function as molecular switches in cells and are key players in intracellular signalling. Ras has been identified as an oncogene and is mutated in more than 20% of human cancers. Here, we report that Bisphenol S binds into a binding pocket of K-Ras4B previously identified for various low molecular weight compounds. Our results advocate for more comprehensive safety studies on the toxicity of Bisphenol S, as it is frequently used for Bisphenol A-free food containers. © 2016 Federation of European Biochemical Societies.

NMR 1H,13C, 15N backbone and 13C side chain resonance assignment of the G12C mutant of human K-Ras bound to GDP.

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Sharma, Alok K; Lee, Seung-Joo; Rigby, Alan C; Townson, Sharon A

2018-05-02

K-Ras is a key driver of oncogenesis, accounting for approximately 80% of Ras-driven human cancers. The small GTPase cycles between an inactive, GDP-bound and an active, GTP-bound state, regulated by guanine nucleotide exchange factors and GTPase activating proteins, respectively. Activated K-Ras regulates cell proliferation, differentiation and survival by signaling through several effector pathways, including Raf-MAPK. Oncogenic mutations that impair the GTPase activity of K-Ras result in a hyperactivated state, leading to uncontrolled cellular proliferation and tumorogenesis. A cysteine mutation at glycine 12 is commonly found in K-Ras associated cancers, and has become a recent focus for therapeutic intervention. We report here 1 H N, 15 N, and 13 C resonance assignments for the 19.3 kDa (aa 1-169) human K-Ras protein harboring an oncogenic G12C mutation in the GDP-bound form (K-RAS G12C-GDP ), using heteronuclear, multidimensional NMR spectroscopy. Backbone 1 H- 15 N correlations have been assigned for all non-proline residues, except for the first methionine residue.

High-density growth arrest in Ras-transformed cells: low Cdk kinase activities in spite of absence of p27^Kip Cdk-complexes

DEFF Research Database (Denmark)

Groth, Anja; Willumsen, Berthe Marie

2005-01-01

The ras oncogene transforms immortalized, contact-inhibited non-malignant murine fibroblasts into cells that are focus forming, exhibit increased saturation density, and are malignant in suitable hosts. Here, we examined changes in cell cycle control complexes as normal and Ras-transformed cells...

Oncogene-induced progression of preneoplastic rat tracheal epithelial cells to neoplasia

International Nuclear Information System (INIS)

Thomassen, D.G.; Kelly, G.

1988-01-01

N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) induced preneoplastic variants of rat tracheal epithelial (RTE) cells can be neo plastically transformed following transfection with oncogenic DNA. Variants differ with respect to the oncogenes required for neoplastic conversion. Polyma virus DNA transformed each of four variants neo plastically, whereas viral ras DNA only transformed two of four variants. These data demonstrate that preneoplastic variants of RTE cells differ with respect to the changes needed for conversion to neoplastic cells and that the variants tested are either at different stages or on different pathways of progression to neoplasia. (author)

Overexpression of K-p21Ras play a prominent role in lung cancer

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Zhang, Peng-bo; Zhou, Xin-liang; Yang, Ju-lun

2018-06-01

The proto-oncogene ras product, p21Ras, has been found overexpression in many human tumors. However, the subtypes of overexpressed p21Ras still remain unclear. The purpose of this study was to investigate overexpressed isoforms of p21Ras and their roles in the progress of lung cancer. Method: The expression of total p21Ras in normal lung tissues and lung cancers was determined by immunohistochemically staining with monoclonal antibody (Mab) KGHR-1 which could recognize and broad spectrum reaction with the (K/H/N) ras protein. Then, the isoforms of p21Ras was examined by specific Mab for each p21Ras subtypes. Results: Low expression of total p21Ras was found in 26.67% (8/30) of normal lung tissues, and 81.31% (87/107) of adenocarcinoma harbored overexpressed total p21Ras. Besides, 70.00% (35/50) of squamous cell carcinoma were detected overexpressed total p21Ras. In addition, 122 lung cancer tissues from overexpression of total p21Ras protein were selected to detect the expression of each subtype. And all the 122 lung cancer tissues were K-p21Ras overexpression. Moreover, there was a statistical significance difference between the expression level of total p21Ras and differentiation, and the same results were observed between the expression level of total p21Ras and lymph node metastasis (P0.05). Conclusions: Overexpression of K-p21Ras plays a prominent role in the progress of lung cancer and it is suggested that the p21Ras could serve as a promising treatment target in lung cancer.

Tumor-Promoting Circuits That Regulate a Cancer-Related Chemokine Cluster: Dominance of Inflammatory Mediators Over Oncogenic Alterations

International Nuclear Information System (INIS)

Leibovich-Rivkin, Tal; Buganim, Yosef; Solomon, Hilla; Meshel, Tsipi; Rotter, Varda; Ben-Baruch, Adit

2012-01-01

Here, we investigated the relative contribution of genetic/signaling components versus microenvironmental factors to the malignancy phenotype. In this system, we took advantage of non-transformed fibroblasts that carried defined oncogenic modifications in Ras and/or p53. These cells were exposed to microenvironmental pressures, and the expression of a cancer-related chemokine cluster was used as readout for the malignancy potential (CCL2, CCL5, CXCL8, CXCL10). In cells kept in-culture, synergism between Ras hyper-activation and p53 dysfunction was required to up-regulate the expression of the chemokine cluster. The in vivo passage of Ras High /p53 Low -modified cells has led to tumor formation, accompanied by potentiation of chemokine release, implicating a powerful role for the tumor microenvironment in up-regulating the chemokine cluster. Indeed, we found that inflammatory mediators which are prevalent in tumor sites, such as TNFα and IL-1β, had a predominant impact on the release of the chemokines, which was substantially higher than that obtained by the oncogenic modifications alone, possibly acting through the transcription factors AP-1 and NF-κB. Together, our results propose that in the unbiased model system that we were using, inflammatory mediators of the tumor milieu have dominating roles over oncogenic modifications in dictating the expression of a pro-malignancy chemokine readout

Ras mutations are rare in solitary cold and toxic thyroid nodules.

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Krohn, K; Reske, A; Ackermann, F; Müller, A; Paschke, R

2001-08-01

Activation of ras proto-oncogenes as a result of point mutations is detectable in a significant percentage of most types of tumour. Similar to neoplasms of other organs, mutations of all three ras genes can be found in thyroid tumours. H-, K- and N-ras mutations have been detected in up to 20% of follicular adenomas and adenomatous nodules which were not functionally characterized. This raises the question as to whether ras mutations are specific for hypofunctional nodules and TSH receptor mutations for hyperfunctioning nodules. To investigate ras and TSH receptor mutations with respect to functional differentiation we studied 41 scintigraphically cold nodules and 47 toxic thyroid nodules. To address the likelihood of a somatic mutation we also studied the clonal origin of these tumours. Genomic DNA was extracted from nodular and surrounding tissue. Mutational hot spots in exons 1 and 2 of the H- and K-ras gene were PCR amplified and sequenced using big dye terminator chemistry. Denaturing gradient gel electrophoresis (DGGE) was used to verify sequencing results for the H-ras gene and to analyse the N-ras gene because its greater sensitivity in detecting somatic mutations. Clonality of nodular thyroid tissue was evaluated using X-Chromosome inactivation based on PCR amplification of the human androgen receptor locus. Monoclonal origin was detectable in 14 of 23 informative samples from cold thyroid nodules. In toxic thyroid nodules the frequency of clonal tissue was 20 in 30 informative cases. Only one point mutation could be found in the N-ras gene codon 61 (Gly to Arg) in a cold adenomatous nodule which was monoclonal. In toxic thyroid nodules no ras mutation was detectable. Our study suggests that ras mutations are rare in solitary cold and toxic thyroid nodules and that the frequent monoclonal origin of these tumours implies somatic mutations in genes other than H-, K- and N-ras.

The oncogenic action of ionizing radiation on rat skin: Progress report, February 1, 1988-January 31, 1989

International Nuclear Information System (INIS)

Burns, F.J.; Garte, S.J.

1988-01-01

Progress is described in 3 general areas corresponding to the specific aims of the proposal, including DNA strand breaks in the epidermis as a function of radiation penetration; oncogene activation in radiation-induced rat skin cancers; and carcinogenesis in rat skin induced by the neon ion beam. Numerous experiments have established that DNA strand breaks per unit dose in the rat epidermis are reduced by about 60% when the radiation penetration is reduced from 1.0 mm to 0.2 mm. The activation of oncogenes in the radiation-induced rat skin cancers followed a pattern. Four highly malignant cancers exhibited activation of K-ras and c-myc oncogenes, while the remaining 8 cancers exhibited only one or the other of these 2 oncogenes. Of 5 squamous carcinomas, 4 showed K-ras activation and 1 showed c-myc activation. Approximately 200 rats were exposed to the neon ion beam at the Bevalac in Berkeley, CA. The carcinogenicity of energetic electrons (2.0 MeV) was determined in conjunction with the neon ion experiment. It is too early to evaluate tumor incidence in the neon ion experiment, but for electrons an unusually large excess of connective tissue tumors, fibromas and sarcomas, have been observed so far. 59 refs., 2 tabs

Eficacia de la terapia génica antisentido utilizando oligonucleótidos anti K-ras y antitelomerasa en cáncer colorrectal

OpenAIRE

Lledó, S.; Alfonso, R.; Aliño, S. F.

2005-01-01

Aim: to test the efficacy of anti-k-ras and antitelomerase oligonucleotides for disabling colorectal cancer cell growth. Material and methods: an established human colorectal cancer cell line (SW 480, ATTC®) was used. Oligodeoxiribonucleotides (ODNs) have a phosphorotioate modification to ensure intracellular intake. We used an antitelomerase ODN (Telp5) and two anti-k-ras ODNs (AS-KRAS and ISIS). AS-KRAS is designed to join the k-ras oncogene's exon 1. ISIS links to the terminal transcriptio...

Interactions of Ras proteins with the plasma membrane and their roles in signaling.

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Eisenberg, Sharon; Henis, Yoav I

2008-01-01

The complex dynamic structure of the plasma membrane plays critical roles in cellular signaling; interactions with the membrane lipid milieu, spatial segregation within and between cellular membranes and/or targeting to specific membrane-associated scaffolds are intimately involved in many signal transduction pathways. In this review, we focus on the membrane interactions of Ras proteins. These small GTPases play central roles in the regulation of cell growth and proliferation, and their excessive activation is commonly encountered in human tumors. Ras proteins associate with the membrane continuously via C-terminal lipidation and additional interactions in both their inactive and active forms; this association, as well as the targeting of specific Ras isoforms to plasma membrane microdomains and to intracellular organelles, have recently been implicated in Ras signaling and oncogenic potential. We discuss biochemical and biophysical evidence for the roles of specific domains of Ras proteins in mediating their association with the plasma membrane, and consider the potential effects of lateral segregation and interactions with membrane-associated protein assemblies on the signaling outcomes.

The Role of Notch Signaling Pathway in Breast Cancer Pathogenesis

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2005-07-01

breast cancer cells, I tested whether ErbB2 overexpression will cooperate with Notch in HMLE cells. While overexpression of activated Notch1 failed to...tyrosine kinase upstream of Ras normally found overexpressed in many breast cancers , also failed to transform HMLE cells. These observations suggested...cooperation between Notch1IC and ErbB2 signaling in transforming HMLE cells. Breast cancers typically do not harbor oncogenic Ras mutations; nevertheless

RAS signaling and anti-RAS therapy: lessons learned from genetically engineered mouse models, human cancer cells, and patient-related studies.

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Fang, Bingliang

2016-01-01

Activating mutations of oncogenic RAS genes are frequently detected in human cancers. The studies in genetically engineered mouse models (GEMMs) reveal that Kras-activating mutations predispose mice to early onset tumors in the lung, pancreas, and gastrointestinal tract. Nevertheless, most of these tumors do not have metastatic phenotypes. Metastasis occurs when tumors acquire additional genetic changes in other cancer driver genes. Studies on clinical specimens also demonstrated that KRAS mutations are present in premalignant tissues and that most of KRAS mutant human cancers have co-mutations in other cancer driver genes, including TP53, STK11, CDKN2A, and KMT2C in lung cancer; APC, TP53, and PIK3CA in colon cancer; and TP53, CDKN2A, SMAD4, and MED12 in pancreatic cancer. Extensive efforts have been devoted to develop therapeutic agents that target enzymes involved in RAS posttranslational modifications, that inhibit downstream effectors of RAS signaling pathways, and that kill RAS mutant cancer cells through synthetic lethality. Recent clinical studies have revealed that sorafenib, a pan-RAF and VEGFR inhibitor, has impressive benefits for KRAS mutant lung cancer patients. Combination therapy of MEK inhibitors with either docetaxel, AKT inhibitors, or PI3K inhibitors also led to improved clinical responses in some KRAS mutant cancer patients. This review discusses knowledge gained from GEMMs, human cancer cells, and patient-related studies on RAS-mediated tumorigenesis and anti-RAS therapy. Emerging evidence demonstrates that RAS mutant cancers are heterogeneous because of the presence of different mutant alleles and/or co-mutations in other cancer driver genes. Effective subclassifications of RAS mutant cancers may be necessary to improve patients' outcomes through personalized precision medicine. © The Author 2015. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology

Real-time single-molecule co-immunoprecipitation analyses reveal cancer-specific Ras signalling dynamics

Science.gov (United States)

Lee, Hong-Won; Kyung, Taeyoon; Yoo, Janghyun; Kim, Tackhoon; Chung, Chaeuk; Ryu, Ji Young; Lee, Hanki; Park, Kihyun; Lee, Sangkyu; Jones, Walton D.; Lim, Dae-Sik; Hyeon, Changbong; Do Heo, Won; Yoon, Tae-Young

2013-01-01

Co-immunoprecipitation (co-IP) has become a standard technique, but its protein-band output provides only static, qualitative information about protein–protein interactions. Here we demonstrate a real-time single-molecule co-IP technique that generates real-time videos of individual protein–protein interactions as they occur in unpurified cell extracts. By analysing single Ras–Raf interactions with a 50-ms time resolution, we have observed transient intermediates of the protein–protein interaction and determined all the essential kinetic rates. Using this technique, we have quantified the active fraction of native Ras proteins in xenograft tumours, normal tissue and cancer cell lines. We demonstrate that the oncogenic Ras mutations selectively increase the active-Ras fraction by one order of magnitude, without affecting total Ras levels or single-molecule signalling kinetics. Our approach allows us to probe the previously hidden, dynamic aspects of weak protein–protein interactions. It also suggests a path forward towards precision molecular diagnostics at the protein–protein interaction level. PMID:23422673

Can K-ras codon 12 mutations be used to distinguish benign bile duct proliferations from metastases in the liver? A molecular analysis of 101 liver lesions from 93 patients

NARCIS (Netherlands)

Hruban, R. H.; Sturm, P. D.; Slebos, R. J.; Wilentz, R. E.; Musler, A. R.; Yeo, C. J.; Sohn, T. A.; van Velthuysen, M. L.; Offerhaus, G. J.

1997-01-01

It can be difficult to distinguish benign bile duct proliferations (BDPs) from well-differentiated metastatic peripancreatic adenocarcinomas on histological grounds alone. Most peripancreatic carcinomas harbor activating point mutations in codon 12 of the K-ras oncogene, suggesting that K-ras

Mechanisms of Membrane Binding of Small GTPase K-Ras4B Farnesylated Hypervariable Region*

Science.gov (United States)

Jang, Hyunbum; Abraham, Sherwin J.; Chavan, Tanmay S.; Hitchinson, Ben; Khavrutskii, Lyuba; Tarasova, Nadya I.; Nussinov, Ruth; Gaponenko, Vadim

2015-01-01

K-Ras4B belongs to a family of small GTPases that regulates cell growth, differentiation and survival. K-ras is frequently mutated in cancer. K-Ras4B association with the plasma membrane through its farnesylated and positively charged C-terminal hypervariable region (HVR) is critical to its oncogenic function. However, the structural mechanisms of membrane association are not fully understood. Here, using confocal microscopy, surface plasmon resonance, and molecular dynamics simulations, we observed that K-Ras4B can be distributed in rigid and loosely packed membrane domains. Its membrane binding domain interaction with phospholipids is driven by membrane fluidity. The farnesyl group spontaneously inserts into the disordered lipid microdomains, whereas the rigid microdomains restrict the farnesyl group penetration. We speculate that the resulting farnesyl protrusion toward the cell interior allows oligomerization of the K-Ras4B membrane binding domain in rigid microdomains. Unlike other Ras isoforms, K-Ras4B HVR contains a single farnesyl modification and positively charged polylysine sequence. The high positive charge not only modulates specific HVR binding to anionic phospholipids but farnesyl membrane orientation. Phosphorylation of Ser-181 prohibits spontaneous farnesyl membrane insertion. The mechanism illuminates the roles of HVR modifications in K-Ras4B targeting microdomains of the plasma membrane and suggests an additional function for HVR in regulation of Ras signaling. PMID:25713064

The Yeast Saccharomyces cerevisiae as a Model for Understanding RAS Proteins and Their Role in Human Tumorigenesis

Science.gov (United States)

Cazzanelli, Giulia; Francisco, Rita; Azevedo, Luísa; Carvalho, Patrícia Dias; Almeida, Ana; Côrte-Real, Manuela; Oliveira, Maria José; Lucas, Cândida; Sousa, Maria João

2018-01-01

The exploitation of the yeast Saccharomyces cerevisiae as a biological model for the investigation of complex molecular processes conserved in multicellular organisms, such as humans, has allowed fundamental biological discoveries. When comparing yeast and human proteins, it is clear that both amino acid sequences and protein functions are often very well conserved. One example of the high degree of conservation between human and yeast proteins is highlighted by the members of the RAS family. Indeed, the study of the signaling pathways regulated by RAS in yeast cells led to the discovery of properties that were often found interchangeable with RAS proto-oncogenes in human pathways, and vice versa. In this work, we performed an updated critical literature review on human and yeast RAS pathways, specifically highlighting the similarities and differences between them. Moreover, we emphasized the contribution of studying yeast RAS pathways for the understanding of human RAS and how this model organism can contribute to unveil the roles of RAS oncoproteins in the regulation of mechanisms important in the tumorigenic process, like autophagy. PMID:29463063

v-Ha-ras oncogene insertion: A model for tumor progression of human small cell lung cancer

International Nuclear Information System (INIS)

Mabry, M.; Nakagawa, Toshitaro; Nelkin, B.D.; McDowell, E.; Gesell, M.; Eggleston, J.C.; Casero, R.A. Jr.; Baylin, S.B.

1988-01-01

Small cell lung cancer (SCLC) manifests a range of phenotypes in culture that may be important in understanding its relationship to non-SCLCs and to tumor progression events in patients. Most SCLC-derived cell lines, termed classic SCLC lines, have properties similar to SCLC tumors in patients. To delineate further the relationships between these phenotypes and the molecular events involved, the authors inserted the v-Ha-ras gene in SCLC cell lines with (biochemical variant) and without (classic) an amplified c-myc gene. These two SCLC subtypes had markedly different phenotypic responses to similar levels of expression of v-Ha-ras RNA. No biochemical or morphologic changes were observed in classic SCLC cells. In contrast, in biochemical variant SCLC cells, v-Ha-ras expression induced features typical of large cell undifferentiated lung carcinoma. Expression of v-Ha-ras in biochemical variant SCLC cells directly demonstrates that important transitions can occur between phenotypes of human lung cancer cells and that these may play a critical role in tumor progression events in patients. The finding provide a model system to study molecular events involved in tumor progression steps within a series of related tumor types

Increased p21ras activity in human fibroblasts transduced with survivin enhances cell proliferation

International Nuclear Information System (INIS)

Temme, Achim; Diestelkoetter-Bachert, Petra; Schmitz, Marc; Morgenroth, Agnieszka; Weigle, Bernd; Rieger, Michael A.; Kiessling, Andrea; Rieber, E. Peter

2005-01-01

Survivin is critically involved in mitosis and when overexpressed enhances the activity of the Aurora B kinase, a serine-threonine kinase belonging to the family of oncogenic Aurora/IpI1p-related kinases. Both proteins interact with Ras GTPase-activating protein suggesting an impact on the Ras pathway. This study aimed at defining the role of survivin in proliferation and potential transformation of cells. When survivin was overexpressed in normal human lung fibroblasts, the characteristic track lanes of fibroblasts were disturbed and the rate of cell proliferation was increased. An enhanced level of p21 ras mRNA and protein expression and concomitant rise in levels of activated p21 ras were observed. Despite increased proliferation cell survival remained dependent on serum and cells were not able to form colonies in soft agar assays. These data suggest that overexpression of survivin increases cell growth but, despite the increase in active p21 ras , is not sufficient to transform primary cells. Yet, in addition to its anti-apoptotic function it might contribute to the accelerated growth of tumour cells by increasing p21 ras activity

Dual knockdown of N-ras and epiregulin synergistically suppressed the growth of human hepatoma cells

Energy Technology Data Exchange (ETDEWEB)

Zhao, Meng; He, Hong-wei; Sun, Huan-xing; Ren, Kai-huan [Department of Oncology, Institute of Medicinal Biotechnology, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing 100050 (China); Shao, Rong-guang, E-mail: shaor@bbn.cn [Department of Oncology, Institute of Medicinal Biotechnology, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing 100050 (China)

2009-09-18

Hepatocellular carcinoma (HCC) is a major challenge because of its resistance to conventional cytotoxic chemotherapy and radiotherapy. Multi-targeted therapy might be a new option for HCC treatment. Our previous study showed that N-ras gene was activated in HCC and was inhibited by RNA interference. In the present study, we investigated the alternation of gene expression by microarray in N-Ras-siRNA-treated HepG2 cells. The results revealed that the EREG gene, encoding epiregulin, was dramatically up-regulated in response to silence of N-ras. We speculated that the up-regulation of epiregulin was involved in the compensatory mechanism of N-ras knockdown for cell growth. Therefore, we evaluated whether dual silence of N-ras and epiregulin display a greater suppression of cell growth. The results confirmed that dual knockdown of N-ras and epiregulin synergistically inhibited cell growth. Our results also showed that dual knockdown of N-ras and epiregulin significantly induced cell arrest at G0/G1 phase. Furthermore, Western blot assay showed that dual knockdown of N-ras and epiregulin markedly reduced the phosphorylations of ERK1/2, Akt and Rb, and inhibited the expression of cyclin D1. Our findings imply that multi-targeted silence of oncogenes might be an effective treatment for HCC.

The Prognostic Impact of K-RAS Mutations in Adult Acute Myeloid Leukemia Patients Treated with High Dose Cytarabine

International Nuclear Information System (INIS)

Ahmad, E.I.; Gawish, H.H.; Al-Azizi, N.M.A.; El-Hefni, A.M.

2009-01-01

Activating point mutation of the RAS gene has been generally accepted as an oncogenic event in a variety of malignancies. It represents one of the most common genetic alterations in acute myeloid leukemia (AML). However there is still controversy about its clinical relevance on the treatment outcome of this leukemia. Objective: This study aimed to clarify the biologic and prognostic impact of K-RAS mutations in relation to the dose of cytarabine (ara-C) used in post induction consolidation chemotherapy in adult AML patients. Patients and Methods: The study comprised 71de novo AML patients with a male: Female ratio of 1.4: 1; their ages ranged from 21-59 years with a median of 37 years. They were subjected to full clinical evaluation, routine laboratory investigations, cytogenetic studies by G banding and K-RAS mutation detection using realtime PCR. The patients were randomized into 2 groups (gps) according to the ara-C dose used in consolidation treatment, HDAC gp receiving 400 mg ara-C and LDAC gp receiving 100 mg ara-C. They were followed over a period of 5 years. Results: Mutations in the K-RAS gene (mutRAS) were detected in 23 patients (32%) with the remaining 48 patients (68%) having wild type RAS (wtRAS). Blast cell percentage was significantly lower in mutRAS compared to wtRAS patients (p=<0.001). The M4 subtype of AML and cases with Inv 16 showed significantly higher frequencies in mutRAS compared to wtRAS patients, (p=0.015, 0.003, respectively). The patients were followed up for a median of 43 months (range 11-57 months). There was no significant difference in overall survival (OS) between mutRAS and wtRAS patients (p=0.326). Within the mutRAS patients treated with HDAC, cumulative OS was significantly higher than those treated with LDAC (p=0.001). This was not the case in the wtRAS group (p=0.285). There was no significant difference in disease The Prognostic Impact of K-RAS Mutations in Adult Acute Myeloid Leukemia Patients Treated with High Dose

DA-Raf, a dominant-negative antagonist of the Ras-ERK pathway, is a putative tumor suppressor.

Science.gov (United States)

Kanno, Emiri; Kawasaki, Osamu; Takahashi, Kazuya; Takano, Kazunori; Endo, Takeshi

2018-01-01

Activating mutations of RAS genes, particularly KRAS, are detected with high frequency in human tumors. Mutated Ras proteins constitutively activate the ERK pathway (Raf-MEK-ERK phosphorylation cascade), leading to cellular transformation and tumorigenesis. DA-Raf1 (DA-Raf) is a splicing variant of A-Raf and contains the Ras-binding domain (RBD) but lacks the kinase domain. Accordingly, DA-Raf antagonizes the Ras-ERK pathway in a dominant-negative fashion and suppresses constitutively activated K-Ras-induced cellular transformation. Thus, we have addressed whether DA-Raf serves as a tumor suppressor of Ras-induced tumorigenesis. DA-Raf(R52Q), which is generated from a single nucleotide polymorphism (SNP) in the RBD, and DA-Raf(R52W), a mutant detected in a lung cancer, neither bound to active K-Ras nor interfered with the activation of the ERK pathway. They were incapable of suppressing activated K-Ras-induced cellular transformation and tumorigenesis in mice, in which K-Ras-transformed cells were transplanted. Furthermore, although DA-Raf was highly expressed in lung alveolar epithelial type 2 (AE2) cells, its expression was silenced in AE2-derived lung adenocarcinoma cell lines with oncogenic KRAS mutations. These results suggest that DA-Raf represents a tumor suppressor protein against Ras-induced tumorigenesis. Copyright © 2017 Elsevier Inc. All rights reserved.

Mechanisms of membrane binding of small GTPase K-Ras4B farnesylated hypervariable region.

Science.gov (United States)

Jang, Hyunbum; Abraham, Sherwin J; Chavan, Tanmay S; Hitchinson, Ben; Khavrutskii, Lyuba; Tarasova, Nadya I; Nussinov, Ruth; Gaponenko, Vadim

2015-04-10

K-Ras4B belongs to a family of small GTPases that regulates cell growth, differentiation and survival. K-ras is frequently mutated in cancer. K-Ras4B association with the plasma membrane through its farnesylated and positively charged C-terminal hypervariable region (HVR) is critical to its oncogenic function. However, the structural mechanisms of membrane association are not fully understood. Here, using confocal microscopy, surface plasmon resonance, and molecular dynamics simulations, we observed that K-Ras4B can be distributed in rigid and loosely packed membrane domains. Its membrane binding domain interaction with phospholipids is driven by membrane fluidity. The farnesyl group spontaneously inserts into the disordered lipid microdomains, whereas the rigid microdomains restrict the farnesyl group penetration. We speculate that the resulting farnesyl protrusion toward the cell interior allows oligomerization of the K-Ras4B membrane binding domain in rigid microdomains. Unlike other Ras isoforms, K-Ras4B HVR contains a single farnesyl modification and positively charged polylysine sequence. The high positive charge not only modulates specific HVR binding to anionic phospholipids but farnesyl membrane orientation. Phosphorylation of Ser-181 prohibits spontaneous farnesyl membrane insertion. The mechanism illuminates the roles of HVR modifications in K-Ras4B targeting microdomains of the plasma membrane and suggests an additional function for HVR in regulation of Ras signaling. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

H-Ras activation promotes cytoplasmic accumulation and phosphoinositide 3-OH kinase association of beta-catenin in epidermal keratinocytes

DEFF Research Database (Denmark)

Espada, J; Pérez-Moreno, M; Braga, V M

1999-01-01

The mechanisms underlying downregulation of the cadherin/catenin complexes and beta-catenin signaling during tumor progression are not fully understood. We have analyzed the effect of oncogenic H-Ras on E-cadherin/catenin complex formation/stabilization and beta-catenin distribution in epidermal ...

Regulation of an H-ras-related transcript by parathyroid hormone in rat osteosarcoma cells

Science.gov (United States)

Scott, D. K.; Weaver, W. R.; Clohisy, J. C.; Brakenhoff, K. D.; Kahn, A. J.; Partridge, N. C.

1992-01-01

The rat osteosarcoma cell line UMR 106-01 is a commonly used model system for the study of osteoblast function. However, it also expresses a phenotype characteristic of transformed cells. To test whether the latter could be accounted for by aberrant oncogene expression, we probed Northern blots of UMR and other osteoblastic cells with a panel of oncogene probes. These blots, when probed with a cDNA specific for v-H-ras, revealed a 7.0-kilobase (kb) H-ras-related transcript (designated HRRT) in UMR 106-01 cells that was not expressed in other osteoblastic cells. Osteoblast-enriched calvarial cells expressed the typical 1.1-kb H-ras mRNA, which was absent in UMR cells. Additionally, Western blots of lysates of UMR cells documented the presence of three proteins immunologically related to H-rasp21. To determine whether HRRT represented a recombinant retrovirus product, Northern blots were probed with a cDNA specific for the highly conserved gag-pol region of Moloney murine leukemia virus. These blots showed parallel cross-reactivity with an apparently identical transcript of 7.0 kb. The 7.0-kb transcripts detected by both v-H-ras and gag-pol probes declined to the same extent after treatment with concentrations of PTH known to inhibit proliferation of these cells. PTH regulated the abundance of HRRT in a time- and dose-dependent manner, with greatest repression of the transcript after 8 h of treatment with 10(-8) M PTH. The decrease in HRRT could not be completely accounted for by changes in transcriptional activity, as determined by nuclear run-on assays.(ABSTRACT TRUNCATED AT 250 WORDS).

Malignant transformation of diploid human fibroblasts by transfection of oncogenes

Energy Technology Data Exchange (ETDEWEB)

McCormick, J.J.

1992-01-01

This document consist of brief reports prepared by postdoctoral students supported by the project, each describing his accomplishments under the grant. Topics include (1) Malignant Transformation of MSU-1. 1 Cells by Gamma Radiation, (2) Correlation between Levels of ras Expression and Presence of Transformed Phenotypes Including Tumorigenicity, Using a Modulatable Promoter, (3) Relation between Specific rad Oncogene Expression, (4) Correlation of Genetic Changes in Fibroblastic Tumors with Malignancies, (5)Transformation of MSU-1.1 Cells by sis Oncogene, (6) Malignant Transformation of MSU-1.0 Cells, (7) Correlation of Urokinase Plasminogen Activation (mu-PA) with Malignant Phenotype, (8)Two Dimensional Gel Electrophoresis Studies of the Proteins of the Major Cell Strains of the MSU-1 Family of Cells, and (9) Correlation between Proteinase Activity Levels and Malignancy.

Malignant transformation of diploid human fibroblasts by transfection of oncogenes

International Nuclear Information System (INIS)

McCormick, J.J.

1992-01-01

This document consist of brief reports prepared by postdoctoral students supported by the project, each describing his accomplishments under the grant. Topics include (1) Malignant Transformation of MSU-1. 1 Cells by Gamma Radiation, (2) Correlation between Levels of ras Expression and Presence of Transformed Phenotypes Including Tumorigenicity, Using a Modulatable Promoter, (3) Relation between Specific rad Oncogene Expression, (4) Correlation of Genetic Changes in Fibroblastic Tumors with Malignancies, (5)Transformation of MSU-1.1 Cells by sis Oncogene, (6) Malignant Transformation of MSU-1.0 Cells, (7) Correlation of Urokinase Plasminogen Activation (mu-PA) with Malignant Phenotype, (8)Two Dimensional Gel Electrophoresis Studies of the Proteins of the Major Cell Strains of the MSU-1 Family of Cells, and (9) Correlation between Proteinase Activity Levels and Malignancy

Mutated N-ras does not induce p19 arf in CO25 cell line | Saleh ...

African Journals Online (AJOL)

The mouse cell line (CO25) used in this study was transfected with a glucocorticoid inducible mutated human N-ras oncogene under transcriptional control of the steroid-sensitive promoter of the mouse mammary tumors virus long terminal repeat MMTV-LTR. This study was aimed to investigate the expression of p19arf and ...

The prognostic impact of K-RAS mutations in adult acute myeloid leukemia patients treated with high-dose cytarabine

Directory of Open Access Journals (Sweden)

Ahmad EI

2011-07-01

Full Text Available Ebtesam I Ahmad, Heba H Gawish, Nashwa MA Al Azizi, Ashraf M ElhefniClinical Pathology Department, Hematology and Oncology Unit of Internal Medicine Department, Faculty of Medicine, Zagazig University, Sharkia, EgyptBackground: Activating point mutation of the RAS gene has been generally accepted as an oncogenic event in a variety of malignancies. It represents one of the most common genetic alterations in acute myeloid leukemia (AML. However, little is known about its clinical relevance in the treatment outcome for this leukemia.Objective: This study aimed to clarify the biologic and prognostic impact of K-RAS mutations in relation to the dose of cytarabine (ara-C used in postinduction consolidation chemotherapy in adult AML patients.Patients and methods: The study comprised of 71 de novo AML patients with male/female ratio 1.4:1; their ages ranged from 21–59 years with a median of 37 years. They were subjected to full clinical evaluation, routine laboratory investigations, cytogenetic studies by G-banding (Giemsa staining, and K-RAS mutation detection using real-time polymerase chain reaction. The patients were randomized into two groups according to the ara-C dose used in consolidation treatment, the high the dose ara-C (HDAC group receiving 400 mg ara-C and-low-dose ara-C (LDAC group receiving 100 mg ara-C; they were followed over a period of five years.Results: Mutations in the K-RAS gene (mutRAS were detected in 23 patients (32% with the remaining 48 patients (68% having wild-type RAS (wtRAS. The percent of blast cells was significantly lower in mutRAS compared to wtRAS patients (P ≤ 0.001 while M4 subtype of AML and Inv(16 frequencies were significantly higher in mutRAS compared to wtRAS patients (P = 0.015 and (P = 0.003, respectively. The patients were followed up for a median of 43 months (range 11–57 months. There was no significant difference in overall survival (OS between mutRAS and wtRAS (P = 0.326. Within the mutRAS

The effect of aquaporin 5 overexpression on the Ras signaling pathway

International Nuclear Information System (INIS)

Woo, Janghee; Lee, Juna; Kim, Myoung Sook; Jang, Se Jin; Sidransky, David; Moon, Chulso

2008-01-01

Human aquaporin 5 (AQP5) has been shown to be overexpressed in multiple cancers, such as pancreatic cancer and colon cancer. Furthermore, it has been reported that ectopic expression of AQP5 leads to many phenotypic changes characteristic of transformation. However, the biochemical mechanism leading to transformation in AQP5-overexpressing cells has not been clearly elucidated. In this report, the overexpression of AQP5 in NIH3T3 cells demonstrated a significant effect on Ras activity and, thus, cell proliferation. Furthermore, this influence was shown to be mediated by phosphorylation of the PKA consensus site of AQP5. This is the first evidence demonstrating an association between AQP5 and a signaling pathway, namely the Ras signal transduction pathway, which may be the basis of the oncogenic properties seen in AQP-overexpressing cells

The APC/C E3 Ligase Complex Activator FZR1 Restricts BRAF Oncogenic Function.

Science.gov (United States)

Wan, Lixin; Chen, Ming; Cao, Juxiang; Dai, Xiangpeng; Yin, Qing; Zhang, Jinfang; Song, Su-Jung; Lu, Ying; Liu, Jing; Inuzuka, Hiroyuki; Katon, Jesse M; Berry, Kelsey; Fung, Jacqueline; Ng, Christopher; Liu, Pengda; Song, Min Sup; Xue, Lian; Bronson, Roderick T; Kirschner, Marc W; Cui, Rutao; Pandolfi, Pier Paolo; Wei, Wenyi

2017-04-01

BRAF drives tumorigenesis by coordinating the activation of the RAS/RAF/MEK/ERK oncogenic signaling cascade. However, upstream pathways governing BRAF kinase activity and protein stability remain undefined. Here, we report that in primary cells with active APC FZR1 , APC FZR1 earmarks BRAF for ubiquitination-mediated proteolysis, whereas in cancer cells with APC-free FZR1, FZR1 suppresses BRAF through disrupting BRAF dimerization. Moreover, we identified FZR1 as a direct target of ERK and CYCLIN D1/CDK4 kinases. Phosphorylation of FZR1 inhibits APC FZR1 , leading to elevation of a cohort of oncogenic APC FZR1 substrates to facilitate melanomagenesis. Importantly, CDK4 and/or BRAF/MEK inhibitors restore APC FZR1 E3 ligase activity, which might be critical for their clinical effects. Furthermore, FZR1 depletion cooperates with AKT hyperactivation to transform primary melanocytes, whereas genetic ablation of Fzr1 synergizes with Pten loss, leading to aberrant coactivation of BRAF/ERK and AKT signaling in mice. Our findings therefore reveal a reciprocal suppression mechanism between FZR1 and BRAF in controlling tumorigenesis. Significance: FZR1 inhibits BRAF oncogenic functions via both APC-dependent proteolysis and APC-independent disruption of BRAF dimers, whereas hyperactivated ERK and CDK4 reciprocally suppress APC FZR1 E3 ligase activity. Aberrancies in this newly defined signaling network might account for BRAF hyperactivation in human cancers, suggesting that targeting CYCLIN D1/CDK4, alone or in combination with BRAF/MEK inhibition, can be an effective anti-melanoma therapy. Cancer Discov; 7(4); 424-41. ©2017 AACR. See related commentary by Zhang and Bollag, p. 356 This article is highlighted in the In This Issue feature, p. 339 . ©2017 American Association for Cancer Research.

H-RAS, K-RAS, and N-RAS gene activation in human bladder cancers.

Science.gov (United States)

Przybojewska, B; Jagiello, A; Jalmuzna, P

2000-08-01

Bladder cancer is one of the leading causes of cancer death in most developed countries. In this work, 19 bladder cancer specimens, along with their infiltrations of the urinary bladder wall from the same patients, were examined for the presence of H-RAS, K-RAS, and N-RAS activation using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. The H-RAS activation was found in 15 (about 84%) of the 19 bladder cancers studied. The same results were obtained in the infiltrating urinary bladder wall samples. N-RAS gene mutations were observed in all cases (except 1) in which H-RAS gene mutations were detected. The results suggest a strong relationship between H-RAS and N-RAS gene activation in bladder cancer. Changes in the K-RAS gene in bladder cancers seem to be a rare event; this is in agreement with findings of other authors. We found activation of the gene in one specimen of bladder cancer and its infiltration of the urinary bladder wall in the same patient.

The Ras effector RASSF2 is a novel tumor-suppressor gene in human colorectal cancer.

Science.gov (United States)

Akino, Kimishige; Toyota, Minoru; Suzuki, Hiromu; Mita, Hiroaki; Sasaki, Yasushi; Ohe-Toyota, Mutsumi; Issa, Jean-Pierre J; Hinoda, Yuji; Imai, Kohzoh; Tokino, Takashi

2005-07-01

Activation of Ras signaling is a hallmark of colorectal cancer (CRC), but the roles of negative regulators of Ras are not fully understood. Our aim was to address that question by surveying genetic and epigenetic alterations of Ras-Ras effector genes in CRC cells. The expression and methylation status of 6 RASSF family genes were examined using RT-PCR and bisulfite PCR in CRC cell lines and in primary CRCs and colorectal adenomas. Colony formation assays and flow cytometry were used to assess the tumor suppressor activities of RASSF1 and RASSF2. Immunofluorescence microscopy was used to determine the effect of altered RASSF2 expression on cell morphology. Mutations of K- ras , BRAF, and p53 were identified using single-strand conformation analysis and direct sequencing. Aberrant methylation and histone deacetylation of RASSF2 was associated with the gene's silencing in CRC. The activities of RASSF2, which were distinct from those of RASSF1, included induction of morphologic changes and apoptosis; moreover, its ability to prevent cell transformation suggests that RASSF2 acts as a tumor suppressor in CRC. Primary CRCs that showed K- ras /BRAF mutations also frequently showed RASSF2 methylation, and inactivation of RASSF2 enhanced K- ras -induced oncogenic transformation. RASSF2 methylation was also frequently identified in colorectal adenomas. RASSF2 is a novel tumor suppressor gene that regulates Ras signaling and plays a pivotal role in the early stages of colorectal tumorigenesis.

Activation of RAS/ERK alone is insufficient to inhibit RXRα function and deplete retinoic acid in hepatocytes

Energy Technology Data Exchange (ETDEWEB)

Wang, Ai-Guo, E-mail: wangaiguotl@hotmail.com; Song, Ya-Nan; Chen, Jun; Li, Hui-Ling; Dong, Jian-Yi; Cui, Hai-Peng; Yao, Liang; Li, Xue-Feng; Gao, Wen-Ting; Qiu, Ze-Wen; Wang, Fu-Jin; Wang, Jing-Yu, E-mail: wangjingyus@163.com

2014-09-26

Highlights: • The activation of RAS/ERK is insufficient to inhibit RXRα function and deplete RA. • The retinoid metabolism-related genes are down-regulated by ras oncogene. • The atRA has no effect on preventing hepatic tumorigenesis or curing the developed hepatic nodules. - Abstract: Activation of RAS/ERK signaling pathway, depletion of retinoid, and phosphorylation of retinoid X receptor alpha (RXRα) are frequent events found in liver tumors and thought to play important roles in hepatic tumorigenesis. However, the relationships among them still remained to be elucidated. By exploring the transgenic mouse model of hepatic tumorigenesis induced by liver-specific expression of H-ras12V oncogene, the activation of RAS/ERK, the mRNA expression levels of retinoid metabolism-related genes, the contents of retinoid metabolites, and phosphorylation of RXRα were determined. RAS/ERK signaling pathway was gradually and significantly activated in hepatic tumor adjacent normal liver tissues (P) and hepatic tumor tissues (T) of H-ras12V transgenic mice compared with normal liver tissues (Wt) of wild type mice. On the contrary, the mRNA expression levels of retinoid metabolism-related genes were significantly reduced in P and T compared with Wt. Interestingly, the retinoid metabolites 9-cis-retinoic acid (9cRA) and all-trans-retinoic acid (atRA), the well known ligands for nuclear transcription factor RXR and retinoic acid receptor (RAR), were significantly decreased only in T compared with Wt and P, although the oxidized polar metabolite of atRA, 4-keto-all-trans-retinoic-acid (4-keto-RA) was significantly decreased in both P and T compared with Wt. To our surprise, the functions of RXRα were significantly blocked only in T compared with Wt and P. Namely, the total protein levels of RXRα were significantly reduced and the phosphorylation levels of RXRα were significantly increased only in T compared with Wt and P. Treatment of H-ras12V transgenic mice at 5-week

K-Ras and β-catenin mutations cooperate with Fgfr3 mutations in mice to promote tumorigenesis in the skin and lung, but not in the bladder

Directory of Open Access Journals (Sweden)

Imran Ahmad

2011-07-01

The human fibroblast growth factor receptor 3 (FGFR3 gene is frequently mutated in superficial urothelial cell carcinoma (UCC. To test the functional significance of FGFR3 activating mutations as a ‘driver’ of UCC, we targeted the expression of mutated Fgfr3 to the murine urothelium using Cre-loxP recombination driven by the uroplakin II promoter. The introduction of the Fgfr3 mutations resulted in no obvious effect on tumorigenesis up to 18 months of age. Furthermore, even when the Fgfr3 mutations were introduced together with K-Ras or β-catenin (Ctnnb1 activating mutations, no urothelial dysplasia or UCC was observed. Interestingly, however, owing to a sporadic ectopic Cre recombinase expression in the skin and lung of these mice, Fgfr3 mutation caused papilloma and promoted lung tumorigenesis in cooperation with K-Ras and β-catenin activation, respectively. These results indicate that activation of FGFR3 can cooperate with other mutations to drive tumorigenesis in a context-dependent manner, and support the hypothesis that activation of FGFR3 signaling contributes to human cancer.

Lipoprotein-biomimetic nanostructure enables efficient targeting delivery of siRNA to Ras-activated glioblastoma cells via macropinocytosis

Science.gov (United States)

Huang, Jia-Lin; Jiang, Gan; Song, Qing-Xiang; Gu, Xiao; Hu, Meng; Wang, Xiao-Lin; Song, Hua-Hua; Chen, Le-Pei; Lin, Ying-Ying; Jiang, Di; Chen, Jun; Feng, Jun-Feng; Qiu, Yong-Ming; Jiang, Ji-Yao; Jiang, Xin-Guo; Chen, Hong-Zhuan; Gao, Xiao-Ling

2017-05-01

Hyperactivated Ras regulates many oncogenic pathways in several malignant human cancers including glioblastoma and it is an attractive target for cancer therapies. Ras activation in cancer cells drives protein internalization via macropinocytosis as a key nutrient-gaining process. By utilizing this unique endocytosis pathway, here we create a biologically inspired nanostructure that can induce cancer cells to `drink drugs' for targeting activating transcription factor-5 (ATF5), an overexpressed anti-apoptotic transcription factor in glioblastoma. Apolipoprotein E3-reconstituted high-density lipoprotein is used to encapsulate the siRNA-loaded calcium phosphate core and facilitate it to penetrate the blood-brain barrier, thus targeting the glioblastoma cells in a macropinocytosis-dependent manner. The nanostructure carrying ATF5 siRNA exerts remarkable RNA-interfering efficiency, increases glioblastoma cell apoptosis and inhibits tumour cell growth both in vitro and in xenograft tumour models. This strategy of targeting the macropinocytosis caused by Ras activation provides a nanoparticle-based approach for precision therapy in glioblastoma and other Ras-activated cancers.

Analysis of beta-catenin, Ki-ras, and microsatellite stability in azoxymethane-induced colon tumors of BDIX/Orl Ico rats

DEFF Research Database (Denmark)

Sørensen, Nanna Møller; Kobaek-Larsen, Morten; Bonne, Anita

2003-01-01

The aim of the study reported here was to investigate whether the azoxymethane (AOM)-induced colon cancer rat model mimics the human situation with regard to microsatellite stability, changes in expression of beta-catenin, and/or changes in the sequence of the proto-oncogene Ki-ras. Colon cancer ...

Cytological and oncogene alterations in radiation-transformed Syrian hamster embryo cells

International Nuclear Information System (INIS)

Trutschler, K.; Hieber, L.; Kellerer, A.M.

1991-01-01

Syrian hamster embryo (SHE) cells were neoplastically transformed by different types of ionizing radiation (γ-rays, α-particles or carbon ions). Transformed and tumor cell lines (derived from nude mice tumors) were analysed for alterations of the oncogenes c-Ha-ras and c-myc, i.e. RFLPs, gene amplifications, activation by point mutation, gene expression, and for cytological changes. In addition, the chromosome number and the numbers of micronuclei per cell have been determined in a series of cell lines. (author)

CREBBP knockdown enhances RAS/RAF/MEK/ERK signaling in Ras pathway mutated acute lymphoblastic leukemia but does not modulate chemotherapeutic response.

Science.gov (United States)

Dixon, Zach A; Nicholson, Lindsay; Zeppetzauer, Martin; Matheson, Elizabeth; Sinclair, Paul; Harrison, Christine J; Irving, Julie A E

2017-04-01

Relapsed acute lymphoblastic leukemia is the most common cause of cancer-related mortality in young people and new therapeutic strategies are needed to improve outcome. Recent studies have shown that heterozygous inactivating mutations in the histone acetyl transferase, CREBBP , are particularly frequent in relapsed childhood acute lymphoblastic leukemia and associated with a hyperdiploid karyotype and KRAS mutations. To study the functional impact of CREBBP haploinsufficiency in acute lymphoblastic leukemia, RNA interference was used to knock down expression of CREBBP in acute lymphoblastic leukemia cell lines and various primagraft acute lymphoblastic leukemia cells. We demonstrate that attenuation of CREBBP results in reduced acetylation of histone 3 lysine 18, but has no significant impact on cAMP-dependent target gene expression. Impaired induction of glucocorticoid receptor targets was only seen in 1 of 4 CREBBP knockdown models, and there was no significant difference in glucocorticoid-induced apoptosis, sensitivity to other acute lymphoblastic leukemia chemotherapeutics or histone deacetylase inhibitors. Importantly, we show that CREBBP directly acetylates KRAS and that CREBBP knockdown enhances signaling of the RAS/RAF/MEK/ERK pathway in Ras pathway mutated acute lymphoblastic leukemia cells, which are still sensitive to MEK inhibitors. Thus, CREBBP mutations might assist in enhancing oncogenic RAS signaling in acute lymphoblastic leukemia but do not alter response to MEK inhibitors. Copyright© Ferrata Storti Foundation.

Status of neutron complex of INR RAS

Energy Technology Data Exchange (ETDEWEB)

Grachev, M I; Koptelov, E A; Kravchuk, L V; Matveev, V A; Perekrestenko, A D; Sidorkin, S F [Institute for Nuclear Research of Russian Academy of Sciences, Prospekt, Moscow (Russian Federation); Stavissky, Y Y

2001-03-01

The neutron complex of INR RAS consists of two sources of neutrons, beam stop, lead slowing down spectrometer and solid state spectrometers. The description of objects and their condition, the program of planned researches, co-operation with other institutes of the Moscow Region, progress reached for last two years are introduced in the article. (author)

Status of neutron complex of INR RAS

International Nuclear Information System (INIS)

Grachev, M.I.; Koptelov, E.A.; Kravchuk, L.V.; Matveev, V.A.; Perekrestenko, A.D.; Sidorkin, S.F.; Stavissky, Y.Y.

2001-01-01

The neutron complex of INR RAS consists of two sources of neutrons, beam stop, lead slowing down spectrometer and solid state spectrometers. The description of objects and their condition, the program of planned researches, co-operation with other institutes of the Moscow Region, progress reached for last two years are introduced in the article. (author)

Syndecan-2 promotes perineural invasion and cooperates with K-ras to induce an invasive pancreatic cancer cell phenotype

Directory of Open Access Journals (Sweden)

De Oliveira Tiago

2012-04-01

Full Text Available Abstract Background We have identified syndecan-2 as a protein potentially involved in perineural invasion of pancreatic adenocarcinoma (PDAC cells. Methods Syndecan-2 (SDC-2 expression was analyzed in human normal pancreas, chronic pancreatitis and PDAC tissues. Functional in vitro assays were carried out to determine its role in invasion, migration and signaling. Results SDC-2 was expressed in the majority of the tested pancreatic cancer cell lines while it was upregulated in nerve-invasive PDAC cell clones. There were 2 distinct expression patterns of SDC-2 in PDAC tissue samples: SDC-2 positivity in the cancer cell cytoplasm and a peritumoral expression. Though SDC-2 silencing (using specific siRNA oligonucleotides did not affect anchorage-dependent growth, it significantly reduced cell motility and invasiveness in the pancreatic cancer cell lines T3M4 and Su8686. On the transcriptional level, migration-and invasion-associated genes were down-regulated following SDC-2 RNAi. Furthermore, SDC-2 silencing reduced K-ras activity, phosphorylation of Src and - further downstream - phosphorylation of ERK2 while levels of the putative SDC-2 signal transducer p120GAP remained unaltered. Conclusion SDC-2 is a novel (perineural invasion-associated gene in PDAC which cooperates with K-ras to induce a more invasive phenotype.

The higher level of complexity of K-Ras4B activation at the membrane.

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Jang, Hyunbum; Banerjee, Avik; Chavan, Tanmay S; Lu, Shaoyong; Zhang, Jian; Gaponenko, Vadim; Nussinov, Ruth

2016-04-01

Is nucleotide exchange sufficient to activate K-Ras4B? To signal, oncogenic rat sarcoma (Ras) anchors in the membrane and recruits effectors by exposing its effector lobe. With the use of NMR and molecular dynamics (MD) simulations, we observed that in solution, farnesylated guanosine 5'-diphosphate (GDP)-bound K-Ras4B is predominantly autoinhibited by its hypervariable region (HVR), whereas the GTP-bound state favors an activated, HVR-released state. On the anionic membrane, the catalytic domain adopts multiple orientations, including parallel (∼180°) and perpendicular (∼90°) alignments of the allosteric helices, with respect to the membrane surface direction. In the autoinhibited state, the HVR is sandwiched between the effector lobe and the membrane; in the active state, with membrane-anchored farnesyl and unrestrained HVR, the catalytic domain fluctuates reinlessly, exposing its effector-binding site. Dimerization and clustering can reduce the fluctuations. This achieves preorganized, productive conformations. Notably, we also observe HVR-autoinhibited K-Ras4B-GTP states, with GDP-bound-like orientations of the helices. Thus, we propose that the GDP/GTP exchange may not be sufficient for activation; instead, our results suggest that the GDP/GTP exchange, HVR sequestration, farnesyl insertion, and orientation/localization of the catalytic domain at the membrane conjointly determine the active or inactive state of K-Ras4B. Importantly, K-Ras4B-GTP can exist in active and inactive states; on its own, GTP binding may not compel K-Ras4B activation.-Jang, H., Banerjee, A., Chavan, T. S, Lu, S., Zhang, J., Gaponenko, V., Nussinov, R. The higher level of complexity of K-Ras4B activation at the membrane. © FASEB.

Activation and amplification of c-Ki-ras in a chemically induced transplantable human pancreas carcinoma

International Nuclear Information System (INIS)

Parsa, I.; Maheshwari, K.K.

1986-01-01

Increasing evidence suggests that carcinogenesis is associated with the stepwise activation of oncogenes. The c-Ki-ras oncogene has been demonstrated in several human solid tumors and is shown to be amplified in tumor cell lines. The authors have probed endonuclease cleaved human pancreas (HP) DNAs and DNAs from an in vitro induced transplantable human pancreas carcinoma (HP-T1) for the presence and/or amplification of c-Ki-ras oncogene. The DNAs were cleaved with BamHI, BgIII, EcoRI, HhaI, HinfI, KpnI, PSTI, PvuII, SaII, SstI, TaqI or XbaI and were subjected to Southern blot analysis using 32 P-labelled EcoRI fragments from HiHi3 clone. The hybridization profiles were similar in both DNAs when digested with BamHI, BgIII, HinfI, KpnI, SaII, SstI, or TaqI. The EcoRI cleaved DNAs from HP and HP-T1 revealed two hybridizing fragments of 6.8 and 3.0 kbp. The 3.0 kbp fragments in DNA from HP-T1 showed more than a 100 folds enhancement as compared to that of HP. The 6.8 hybridizing fragments also appeared 10 fold greater in HP-T1 DNA. Similar enhancements were also present in HP-T1 DNA cleaved with PstI and PvuII. Preliminary results from comparison of poly(A) + RNAs, prepared from total HP and HP-T1 RNAs, by Northern blot analysis using the same probe reflect similar enhancement in RNA from transplantable pancreas carcinoma

Prognostic implications of c-Ki-ras2 mutations in patients with advanced colorectal cancer treated with 5-fluorouracil and interferon: a study of the eastern cooperative oncology group (EST 2292)

Science.gov (United States)

Wadler, S; Bajaj, R; Neuberg, D; Agarwal, V; Haynes, H; Benson, A B

1997-01-01

Mutations in c-Ki-ras2 (ras) occur in about 40% of patients with colorectal cancers and occur early in the pathogenesis of this disease. To evaluate the prognostic value of mutations in ras, the Eastern Cooperative Oncology Group (ECOG) conducted a retrospective study (EST 2292) to determine the frequency of mutations in patients with advanced colorectal cancer, and to determine whether ras mutations were associated with altered response to therapy and survival. Patients were enrolled from four studies: P-Z289, an ECOG phase II trial of 5-fluorouracil (5-FU) and interferon (IFN) in patients with advanced colorectal cancer; P-Z991, an ECOG phase I trial of 5-FU and IFN in patients with advanced malignancies; and two trials from the Albert Einstein College of Medicine in patients with advanced colorectal cancer treated with 5-FU and either IFN-alpha or IFN-beta. All patients had advanced colorectal carcinoma and had sufficient histologic material available for analysis for the presence and type of ras, using polymerase chain reaction and dot-blot analysis with sets of probes sufficient to detect all the common mutations of ras at codons 12, 13, and 61. Seventy-two patients were enrolled in this trial. Mutations in ras were detected in 25 (35%), including 17 (23%) in codon 12, four (6%) in codon 13, and four (6%) in codon 61. There was no correlation between the presence of a ras mutation and age, sex, Dukes' stage, histology, or tumor markers. Thirty-one of 72 patients (43%) responded to therapy with 5-FU and IFN, and 10 of 31 responders (32%) and 15 of 41 nonresponders (37%) had mutations in ras. There was no difference in response rates or overall survival between the groups with and without ras mutations. It is unlikely that ras mutations will have significant prognostic value for either response to therapy or survival in patients with colorectal carcinomas treated with 5-FU and IFN.

The higher level of complexity of K-Ras4B activation at the membrane

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Jang, Hyunbum; Banerjee, Avik; Chavan, Tanmay S.; Lu, Shaoyong; Zhang, Jian; Gaponenko, Vadim; Nussinov, Ruth

2016-01-01

Is nucleotide exchange sufficient to activate K-Ras4B? To signal, oncogenic rat sarcoma (Ras) anchors in the membrane and recruits effectors by exposing its effector lobe. With the use of NMR and molecular dynamics (MD) simulations, we observed that in solution, farnesylated guanosine 5′-diphosphate (GDP)-bound K-Ras4B is predominantly autoinhibited by its hypervariable region (HVR), whereas the GTP-bound state favors an activated, HVR-released state. On the anionic membrane, the catalytic domain adopts multiple orientations, including parallel (∼180°) and perpendicular (∼90°) alignments of the allosteric helices, with respect to the membrane surface direction. In the autoinhibited state, the HVR is sandwiched between the effector lobe and the membrane; in the active state, with membrane-anchored farnesyl and unrestrained HVR, the catalytic domain fluctuates reinlessly, exposing its effector-binding site. Dimerization and clustering can reduce the fluctuations. This achieves preorganized, productive conformations. Notably, we also observe HVR-autoinhibited K-Ras4B-GTP states, with GDP-bound-like orientations of the helices. Thus, we propose that the GDP/GTP exchange may not be sufficient for activation; instead, our results suggest that the GDP/GTP exchange, HVR sequestration, farnesyl insertion, and orientation/localization of the catalytic domain at the membrane conjointly determine the active or inactive state of K-Ras4B. Importantly, K-Ras4B-GTP can exist in active and inactive states; on its own, GTP binding may not compel K-Ras4B activation.—Jang, H., Banerjee, A., Chavan, T. S, Lu, S., Zhang, J., Gaponenko, V., Nussinov, R. The higher level of complexity of K-Ras4B activation at the membrane. PMID:26718888

Which downstream signal transduction pathway(s) of H-ras are necessary for the cellular response(s) to ionizing radiation? (Results of an astro research fellowship year)

International Nuclear Information System (INIS)

Rudoltz, Marc S.; Muschel, Ruth J.; McKenna, W. Gillies

1996-01-01

Purpose/Background: The H-ras oncogene encodes a protein which is an essential component of multiple downstream effector pathways required for induction of proliferation and differentiation. Ras plays a role in the control some of these signal transduction pathways, such as the MAP kinase pathway which controls gene expression and the Rac-Rho pathway which controls cell morphology. Previous work from our laboratory has associated H-ras expression with radiation resistance, a prolonged delay in G2 following exposure to ionizing radiation, and suppression of radiation-induced apoptosis. In addition, H-ras cooperates with myc in transformation. Recent work by White et al. (Cell 80:533-541, 1995) and Joneson et al. (Science 271: 810-812, 1996) describes three mutations in H-ras which were engineered to eliminate different downstream signal transduction pathways of H-ras. T35S contains a serine in place of threonine at amino acid 35 and is defective for ras-induced cytoskeletal changes and initiation of DNA synthesis. E37G contains a glutamic acid in place of glycine at amino acid 37 which eliminates interaction of H-ras with a GDP/GTP exchange factor. C40 contains a substitution of cysteine for tyrosine at amino acid 40 and is defective for H-ras induction of the MAP kinase pathway. We propose that by expressing these mutant H-ras proteins in immortalized cells the downstream pathways of H-ras which regulate the cellular response(s) to ionizing radiation may be determined. Materials and Methods: pHP-5 plasmids encoding these H-ras mutant genes (see White et al.) were transfected by calcium phosphate precipitation into MR4 cells, rat embryo fibroblasts immortalized by expression of v-myc. In this vector, the cDNA for H-ras is placed under the control of a CMV constitutive promoter, and selection is provided by hygromycin. The transfections performed were as follows: V12Ras (no mutation), T35S, E37G, C40, T35S + E37G, and T35S + C40. Twenty four hours after transfection

Activation of the JNK pathway is essential for transformation by the Met oncogene.

Science.gov (United States)

Rodrigues, G A; Park, M; Schlessinger, J

1997-05-15

The Met/Hepatocyte Growth Factor (HGF) receptor tyrosine kinase is oncogenically activated through a rearrangement that creates a hybrid gene Tpr-Met. The resultant chimeric p65(Tpr-Met) protein is constitutively phosphorylated on tyrosine residues in vivo and associates with a number of SH2-containing signaling molecules including the p85 subunit of PI-3 kinase and the Grb2 adaptor protein, which couples receptor tyrosine kinases to the Ras signaling pathway. Mutation of the binding site for Grb2 impairs the ability of Tpr-Met oncoprotein to transform fibroblasts, suggesting that the activation of the Ras/MAP kinase signaling pathway through Grb2 may be essential for cellular transformation. To test this hypothesis dominant-negative mutants of Grb2 with deletions of the SH3 domains were introduced into Tpr-Met transformed fibroblasts. Cells overexpressing the mutants were found to be morphologically reverted and exhibited reduced growth in soft agar. Surprisingly, the Grb2 mutants blocked activation of the JNK/SAPK but not MAP kinase activity induced by the Tpr-Met oncoprotein. Additionally, cells expressing dominant-negative Grb2 mutants had reduced PI-3-kinase activity and dominant-negative mutants of Rac1 blocked both Tpr-Met-induced transformation and activation of JNK. These experiments reveal a novel link between Met and the JNK pathway, which is essential for transformation by this oncogene.

The Ras suppressor-1 (RSU-1 in cancer

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Lefteris C Zacharia

2017-04-01

Full Text Available Primary tumors are seldom the cause of death for cancer patients as most patients die from metastatic disease. Thus, deciphering metastatic mechanisms and key molecules involved is of utmost importance for the improved survival of cancer patients. Metastasis is a complex process in which cancer cells dissociate from the original tumor and spread to distant sites of the body. During the metastatic process, cancer cells lose contact both with the extracellular matrix (ECM and the neighboring cells within the primary tumor, thus invading though surrounding tissues. Therefore, ECM, and ECM-related adhesion proteins play a critical role in the metastatic process. Ras suppressor-1 (RSU-1 was first identified as a suppressor of Ras-dependent oncogenic transformation and is localized to cell-ECM adhesions where it is known to interact with the pro-survival adhesion protein PINCH-1. Although the connection to cancer is obvious, little is known regarding its expression in various cancer types. This opinion piece is focusing on recent literature regarding the expression of RSU-1 in various cancer types and the possible molecular mechanism of its action, pointing towards questions that need still to be addressed in this research field.

Characterization of TRPS1 and ERAS as oncogenes implicated in breast cancer

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Lopez Gonzalez, L.

2015-07-01

New high throughput technologies have made possible to identify putative oncogenes in breast cancer. In this project we aim to relate and characterise two novel putative oncogenes, ERAS and TRPS1, in their role in human breast cancer. TRPS1, an atypical GATA factor, modulates cell proliferation and controls cell cycle progression through repression of GATA-regulated genes, therefore acting as a tumour suppressor gene. Conversely, TRPS1 expression has been shown to be significantly elevated in luminal and in a lesser extent in basal breast cancer cells, presenting roles both as an oncogene and as a tumour suppressor gene in breast cancer development. The aim of this project is therefore to determine the characteristics of TRPS1 either as a putative novel human oncogene or as a tumour suppressor gene in breast cancer cells. To this aim, we have cloned a novel isoform of TRPS1 and introduced it into several breast cancer cell lines. Our results show that overexpression of this isoform of TRPS1 results in variations in motility in non-carcinogenic cell lines, as well as in a series of EMT-like changes such as the down-regulation of the EMT marker E-cadherin, both of which can be associated to an increase in malignancy, suggesting an oncogenic behaviour for TRPS1. Furthermore, our results suggest that constitutively active members of the RAS protein family induce the expression of TRPS1, establishing a relationship between both genes. We can conclude that the effects of TRPS1 overexpression are moderate, inducing some changes but not fully transforming the cells. Therefore we cannot confirm that TRPS1 is a putative oncogene in breast cancer. (Author)

Cytogenetic characterization and H-ras associated transformation of immortalized human mammary epithelial cells

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Larivee Siobhan

2006-05-01

Full Text Available Abstract Introduction Immortalization is a key step in malignant transformation, but immortalization alone is insufficient for transformation. Human mammary epithelial cell (HMEC transformation is a complex process that requires additional genetic changes beyond immortalization and can be accomplished in vitro by accumulation of genetic changes and expression of H-ras. Methods HMEC were immortalized by serial passaging and transduction with the catalytic subunit of the human telomerase gene (hTERT. The immortalized cells were passaged in vitro and studied by a combination of G- banding and Spectral Karyotyping (SKY. H-ras transduced, hTERT immortalized cells were cloned in soft agar and injected into nude mice. Extensive analysis was performed on the tumors that developed in nude mice, including immunohistochemistry and western blotting. Results Immortal HMEC alone were not tumorigenic in γ-irradiated nude mice and could not grow in soft agar. Late passage hTERT immortalized HMEC from a donor transduced with a retroviral vector containing the mutant, autoactive, human H-ras61L gene acquired anchorage independent growth properties and the capacity for tumorigenic growth in vivo. The tumors that developed in the nude mice were poorly differentiated epithelial carcinomas that continued to overexpress ras. These cells were resistant to doxorubicin mediated G1/S phase arrest but were sensitive to treatment with a farnesyltransferase inhibitor. Conclusion Some of the cytogenetic changes are similar to what is observed in premalignant and malignant breast lesions. Despite these changes, late passage immortal HMEC are not tumorigenic and could only be transformed with overexpression of a mutant H-ras oncogene.

A Drosophila immune response against Ras-induced overgrowth

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Thomas Hauling

2014-03-01

Full Text Available Our goal is to characterize the innate immune response against the early stage of tumor development. For this, animal models where genetic changes in specific cells and tissues can be performed in a controlled way have become increasingly important, including the fruitfly Drosophila melanogaster. Many tumor mutants in Drosophila affect the germline and, as a consequence, also the immune system itself, making it difficult to ascribe their phenotype to a specific tissue. Only during the past decade, mutations have been induced systematically in somatic cells to study the control of tumorous growth by neighboring cells and by immune cells. Here we show that upon ectopic expression of a dominant-active form of the Ras oncogene (RasV12, both imaginal discs and salivary glands are affected. Particularly, the glands increase in size, express metalloproteinases and display apoptotic markers. This leads to a strong cellular response, which has many hallmarks of the granuloma-like encapsulation reaction, usually mounted by the insect against larger foreign objects. RNA sequencing of the fat body reveals a characteristic humoral immune response. In addition we also identify genes that are specifically induced upon expression of RasV12. As a proof-of-principle, we show that one of the induced genes (santa-maria, which encodes a scavenger receptor, modulates damage to the salivary glands. The list of genes we have identified provides a rich source for further functional characterization. Our hope is that this will lead to a better understanding of the earliest stage of innate immune responses against tumors with implications for mammalian immunity.

Molecular markers for diagnostic cytology of neoplasms in the head region of the pancreas: mutation of K-ras and overexpression of the p53 protein product

NARCIS (Netherlands)

van Es, J. M.; Polak, M. M.; van den Berg, F. M.; Ramsoekh, T. B.; Craanen, M. E.; Hruban, R. H.; Offerhaus, G. J.

1995-01-01

To determine the potential efficiency of molecular markers specific for neoplastic change--mutations of the K-ras oncogene and the p53 tumour suppressor gene--in diagnosing pancreatic carcinoma. Archival cytology samples obtained from 17 patients with established pancreatic carcinoma were assayed

A high level of liver-specific expression of oncogenic KrasV12 drives robust liver tumorigenesis in transgenic zebrafish

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Anh Tuan Nguyen

2011-11-01

Human liver cancer is one of the deadliest cancers worldwide, with hepatocellular carcinoma (HCC being the most common type. Aberrant Ras signaling has been implicated in the development and progression of human HCC, but a complete understanding of the molecular mechanisms of this protein in hepatocarcinogenesis remains elusive. In this study, a stable in vivo liver cancer model using transgenic zebrafish was generated to elucidate Ras-driven tumorigenesis in HCC. Using the liver-specific fabp10 (fatty acid binding protein 10 promoter, we overexpressed oncogenic krasV12 specifically in the transgenic zebrafish liver. Only a high level of krasV12 expression initiated liver tumorigenesis, which progressed from hyperplasia to benign and malignant tumors with activation of the Ras-Raf-MEK-ERK and Wnt–β-catenin pathways. Histological diagnosis of zebrafish tumors identified HCC as the main lesion. The tumors were invasive and transplantable, indicating malignancy of these HCC cells. Oncogenic krasV12 was also found to trigger p53-dependent senescence as a tumor suppressive barrier in the pre-neoplastic stage. Microarray analysis of zebrafish liver hyperplasia and HCC uncovered the deregulation of several stage-specific and common biological processes and signaling pathways responsible for krasV12-driven liver tumorigenesis that recapitulated the molecular hallmarks of human liver cancer. Cross-species comparisons of cancer transcriptomes further defined a HCC-specific gene signature as well as a liver cancer progression gene signature that are evolutionarily conserved between human and zebrafish. Collectively, our study presents a comprehensive portrait of molecular mechanisms during progressive Ras-induced HCC. These observations indicate the validity of our transgenic zebrafish to model human liver cancer, and this model might act as a useful platform for drug screening and identifying new therapeutic targets.

Multifunctional imaging signature for V-KI-RAS2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations in colorectal cancer.

Science.gov (United States)

Miles, Kenneth A; Ganeshan, Balaji; Rodriguez-Justo, Manuel; Goh, Vicky J; Ziauddin, Zia; Engledow, Alec; Meagher, Marie; Endozo, Raymondo; Taylor, Stuart A; Halligan, Stephen; Ell, Peter J; Groves, Ashley M

2014-03-01

This study explores the potential for multifunctional imaging to provide a signature for V-KI-RAS2 Kirsten rat sarcoma viral oncogene homolog (KRAS) gene mutations in colorectal cancer. This prospective study approved by the institutional review board comprised 33 patients undergoing PET/CT before surgery for proven primary colorectal cancer. Tumor tissue was examined histologically for presence of the KRAS mutations and for expression of hypoxia-inducible factor-1 (HIF-1) and minichromosome maintenance protein 2 (mcm2). The following imaging parameters were derived for each tumor: (18)F-FDG uptake ((18)F-FDG maximum standardized uptake value [SUVmax]), CT texture (expressed as mean of positive pixels [MPP]), and blood flow measured by dynamic contrast-enhanced CT. A recursive decision tree was developed in which the imaging investigations were applied sequentially to identify tumors with KRAS mutations. Monte Carlo analysis provided mean values and 95% confidence intervals for sensitivity, specificity, and accuracy. The final decision tree comprised 4 decision nodes and 5 terminal nodes, 2 of which identified KRAS mutants. The true-positive rate, false-positive rate, and accuracy (95% confidence intervals) of the decision tree were 82.4% (63.9%-93.9%), 0% (0%-10.4%), and 90.1% (79.2%-96.0%), respectively. KRAS mutants with high (18)F-FDG SUVmax and low MPP showed greater frequency of HIF-1 expression (P = 0.032). KRAS mutants with low (18)F-FDG SUV(max), high MPP, and high blood flow expressed mcm2 (P = 0.036). Multifunctional imaging with PET/CT and recursive decision-tree analysis to combine measurements of tumor (18)F-FDG uptake, CT texture, and perfusion has the potential to identify imaging signatures for colorectal cancers with KRAS mutations exhibiting hypoxic or proliferative phenotypes.

Depletion of Pokemon gene inhibits hepatocellular carcinoma cell growth through inhibition of H-ras.

Science.gov (United States)

Zhang, Quan-Le; Tian, De-An; Xu, Xiang-Jiang

2011-01-01

Pokemon is a transcription repressor which plays a critical role in cell transformation and malignancy. However, little is known about its effect on the development and progression of hepatocellular carcinoma (HCC). The aim of this study was to investigate the expression of Pokemon in human HCC tissues and the biological behavior of Pokemon in HCC cells in which it is overexpressed. We also explored the expression of potential downstream cofactors of Pokemon. Reverse transcription polymerase chain reaction and Western blot analysis were used to investigate the expression of Pokemon in tissues of 30 HCC patients. We then examined cell proliferation or apoptosis and β-catenin or H-ras expression in Pokemon-depleted HepG(2) cells using DNA vector-based RNA interference technology. Pokemon was markedly expressed in 22/30 (73.3%) HCC tissues, with expression levels higher than in adjacent normal liver tissues (p Pokemon inhibited proliferation of HepG(2) or induced apoptosis. Also, H-ras expression decreased to a large extent. Pokemon exerts its oncogenic activity in the development of HCC by promoting cancer cell growth and reducing apoptosis, and the effect may be mediated by H-ras. Copyright © 2011 S. Karger AG, Basel.

Evolution of AF6-RAS association and its implications in mixed-lineage leukemia

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Smith, Matthew J.; Ottoni, Elizabeth; Ishiyama, Noboru; Goudreault, Marilyn; Haman, André; Meyer, Claus; Tucholska, Monika; Gasmi-Seabrook, Genevieve; Menezes, Serena; Laister, Rob C.; Minden, Mark D.; Marschalek, Rolf; Gingras, Anne-Claude; Hoang, Trang; Ikura, Mitsuhiko

2017-10-23

Elucidation of activation mechanisms governing protein fusions is essential for therapeutic development. MLL undergoes rearrangement with numerous partners, including a recurrent translocation fusing the epigenetic regulator to a cytoplasmic RAS effector, AF6/afadin. We show here that AF6 employs a non-canonical, evolutionarily conserved α-helix to bind RAS, unique to AF6 and the classical RASSF effectors. Further, all patients with MLL-AF6 translocations express fusion proteins missing only this helix from AF6, resulting in exposure of hydrophobic residues that induce dimerization. We provide evidence that oligomerization is the dominant mechanism driving oncogenesis from rare MLL translocation partners and employ our mechanistic understanding of MLL-AF6 to examine how dimers induce leukemia. Proteomic data resolve association of dimerized MLL with gene expression modulators, and inhibiting dimerization disrupts formation of these complexes while completely abrogating leukemogenesis in mice. Oncogenic gene translocations are thus selected under pressure from protein structure/function, underscoring the complex nature of chromosomal rearrangements.

The RAS Initiative

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NCI established the RAS Initiative to explore innovative approaches for attacking the proteins encoded by mutant forms of RAS genes and to ultimately create effective, new therapies for RAS-related cancers.

R-Ras regulates migration through an interaction with filamin A in melanoma cells.

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Joanna E Gawecka

2010-06-01

Full Text Available Changes in cell adhesion and migration in the tumor microenvironment are key in the initiation and progression of metastasis. R-Ras is one of several small GTPases that regulate cell adhesion and migration on the extracellular matrix, however the mechanism has not been completely elucidated. Using a yeast two-hybrid approach we sought to identify novel R-Ras binding proteins that might mediate its effects on integrins.We identified Filamin A (FLNa as a candidate interacting protein. FLNa is an actin-binding scaffold protein that also binds to integrin beta1, beta2 and beta7 tails and is associated with diverse cell processes including cell migration. Indeed, M2 melanoma cells require FLNa for motility. We further show that R-Ras and FLNa interact in co-immunoprecipitations and pull-down assays. Deletion of FLNa repeat 3 (FLNaDelta3 abrogated this interaction. In M2 melanoma cells active R-Ras co-localized with FLNa but did not co-localize with FLNa lacking repeat 3. Thus, activated R-Ras binds repeat 3 of FLNa. The functional consequence of this interaction was that active R-Ras and FLNa coordinately increased cell migration. In contrast, co-expression of R-Ras and FLNaDelta3 had a significantly reduced effect on migration. While there was enhancement of integrin activation and fibronectin matrix assembly, cell adhesion was not altered. Finally, siRNA knockdown of endogenous R-Ras impaired FLNa-dependent fibronectin matrix assembly.These data support a model in which R-Ras functionally associates with FLNa and thereby regulates integrin-dependent migration. Thus in melanoma cells R-Ras and FLNa may cooperatively promote metastasis by enhancing cell migration.

Interaction between Wnt/β-catenin and RAS-ERK pathways and an anti-cancer strategy via degradations of β-catenin and RAS by targeting the Wnt/β-catenin pathway.

Science.gov (United States)

Jeong, Woo-Jeong; Ro, Eun Ji; Choi, Kang-Yell

2018-01-01

Aberrant activation of the Wnt/β-catenin and RAS-extracellular signal-regulated kinase (ERK) pathways play important roles in the tumorigenesis of many different types of cancer, most notably colorectal cancer (CRC). Genes for these two pathways, such as adenomatous polyposis coli ( APC ) and KRAS are frequently mutated in human CRC, and involved in the initiation and progression of the tumorigenesis, respectively. Moreover, recent studies revealed interaction of APC and KRAS mutations in the various stages of colorectal tumorigenesis and even in metastasis accompanying activation of the cancer stem cells (CSCs). A key event in the synergistic cooperation between Wnt/β-catenin and RAS-ERK pathways is a stabilization of both β-catenin and RAS especially mutant KRAS by APC loss, and pathological significance of this was indicated by correlation of increased β-catenin and RAS levels in human CRC where APC mutations occur as high as 90% of CRC patients. Together with the notion of the protein activity reduction by lowering its level, inhibition of both β-catenin and RAS especially by degradation could be a new ideal strategy for development of anti-cancer drugs for CRC. In this review, we will discuss interaction between the Wnt/β-catenin and RAS-ERK pathways in the colorectal tumorigenesis by providing the mechanism of RAS stabilization by aberrant activation of Wnt/β-catenin. We will also discuss our small molecular anti-cancer approach controlling CRC by induction of specific degradations of both β-catenin and RAS via targeting Wnt/β-catenin pathway especially for the KYA1797K, a small molecule specifically binding at the regulator of G-protein signaling (RGS)-domain of Axin.

High-throughput screening identifies small molecules that bind to the RAS:SOS:RAS complex and perturb RAS signaling.

Science.gov (United States)

Burns, Michael C; Howes, Jennifer E; Sun, Qi; Little, Andrew J; Camper, DeMarco V; Abbott, Jason R; Phan, Jason; Lee, Taekyu; Waterson, Alex G; Rossanese, Olivia W; Fesik, Stephen W

2018-05-01

K-RAS is mutated in approximately 30% of human cancers, resulting in increased RAS signaling and tumor growth. Thus, RAS is a highly validated therapeutic target, especially in tumors of the pancreas, lung and colon. Although directly targeting RAS has proven to be challenging, it may be possible to target other proteins involved in RAS signaling, such as the guanine nucleotide exchange factor Son of Sevenless (SOS). We have previously reported on the discovery of small molecules that bind to SOS1, activate SOS-mediated nucleotide exchange on RAS, and paradoxically inhibit ERK phosphorylation (Burns et al., PNAS, 2014). Here, we describe the discovery of additional, structurally diverse small molecules that also bind to SOS1 in the same pocket and elicit similar biological effects. We tested >160,000 compounds in a fluorescence-based assay to assess their effects on SOS-mediated nucleotide exchange. X-Ray structures revealed that these small molecules bind to the CDC25 domain of SOS1. Compounds that elicited high levels of nucleotide exchange activity in vitro increased RAS-GTP levels in cells, and inhibited phospho ERK levels at higher treatment concentrations. The identification of structurally diverse SOS1 binding ligands may assist in the discovery of new molecules designed to target RAS-driven tumors. Copyright © 2018 Elsevier Inc. All rights reserved.

PIK3CA mutations frequently coexist with RAS and BRAF mutations in patients with advanced cancers.

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Filip Janku

Full Text Available Oncogenic mutations of PIK3CA, RAS (KRAS, NRAS, and BRAF have been identified in various malignancies, and activate the PI3K/AKT/mTOR and RAS/RAF/MEK pathways, respectively. Both pathways are critical drivers of tumorigenesis.Tumor tissues from 504 patients with diverse cancers referred to the Clinical Center for Targeted Therapy at MD Anderson Cancer Center starting in October 2008 were analyzed for PIK3CA, RAS (KRAS, NRAS, and BRAF mutations using polymerase chain reaction-based DNA sequencing.PIK3CA mutations were found in 54 (11% of 504 patients tested; KRAS in 69 (19% of 367; NRAS in 19 (8% of 225; and BRAF in 31 (9% of 361 patients. PIK3CA mutations were most frequent in squamous cervical (5/14, 36%, uterine (7/28, 25%, breast (6/29, 21%, and colorectal cancers (18/105, 17%; KRAS in pancreatic (5/9, 56%, colorectal (49/97, 51%, and uterine cancers (3/20, 15%; NRAS in melanoma (12/40, 30%, and uterine cancer (2/11, 18%; BRAF in melanoma (23/52, 44%, and colorectal cancer (5/88, 6%. Regardless of histology, KRAS mutations were found in 38% of patients with PIK3CA mutations compared to 16% of patients with wild-type (wtPIK3CA (p = 0.001. In total, RAS (KRAS, NRAS or BRAF mutations were found in 47% of patients with PIK3CA mutations vs. 24% of patients wtPIK3CA (p = 0.001. PIK3CA mutations were found in 28% of patients with KRAS mutations compared to 10% with wtKRAS (p = 0.001 and in 20% of patients with RAS (KRAS, NRAS or BRAF mutations compared to 8% with wtRAS (KRAS, NRAS or wtBRAF (p = 0.001.PIK3CA, RAS (KRAS, NRAS, and BRAF mutations are frequent in diverse tumors. In a wide variety of tumors, PIK3CA mutations coexist with RAS (KRAS, NRAS and BRAF mutations.

Utilizing ras signaling pathway to direct selective replication of herpes simplex virus-1.

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Weihong Pan

Full Text Available Re-engineering the tropism of viruses is an attractive translational strategy for targeting cancer cells. The Ras signal transduction pathway is a central hub for a variety of pro-oncogenic events with a fundamental role in normal and neoplastic physiology. In this work we were interested in linking Ras activation to HSV-1 replication in a direct manner in order to generate a novel oncolytic herpes virus which can target cancer cells. To establish such link, we developed a mutant HSV-1 in which the expression of ICP4 (infected cell protein-4, a viral protein necessary for replication is controlled by activation of ELK, a transcription factor down-stream of the Ras pathway and mainly activated by ERK (extracellular signal-regulated kinase, an important Ras effector pathway. This mutant HSV-1 was named as Signal-Smart 1 (SS1. A series of prostate cells were infected with the SS1 virus. Cells with elevated levels of ELK activation were preferentially infected by the SS1 virus, as demonstrated by increased levels of viral progeny, herpetic glycoprotein C and overall SS1 viral protein production. Upon exposure to SS1, the proliferation, invasiveness and colony formation capabilities of prostate cancer cells with increased ELK activation were significantly decreased (p<0.05, while the rate of apoptosis/necrosis in these cells was increased. Additionally, high Ras signaling cells infected with SS1 showed a prominent arrest in the G1 phase of the cell cycle as compared to cells exposed to parental HSV-1. The results of this study reveal the potential for re-modeling the host-herpes interaction to specifically interfere with the life of cancer cells with increased Ras signaling. SS1 also serves as a "prototype" for development of a family of signal-smart viruses which can target cancer cells on the basis of their signaling portfolio.

Oxidative Stress Posttranslationally Regulates the Expression of Ha-Ras and Ki-Ras in Cultured Astrocytes

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Samantha Messina

2012-01-01

Full Text Available Addition of hydrogen peroxide to cultured astrocytes induced a rapid and transient increase in the expression of Ha-Ras and Ki-Ras. Pull-down experiments with the GTP-Ras-binding domain of Raf-1 showed that oxidative stress substantially increased the activation of Ha-Ras, whereas a putative farnesylated activated form of Ki-Ras was only slightly increased. The increase in both Ha-Ras and Ki-Ras was insensitive to the protein synthesis inhibitor, cycloheximide, and was occluded by the proteasomal inhibitor, MG-132. In addition, exposure to hydrogen peroxide reduced the levels of ubiquitinated Ras protein, indicating that oxidative stress leads to a reduced degradation of both isoforms through the ubiquitin/proteasome pathway. Indeed, the late reduction in Ha-Ras and Ki-Ras was due to a recovery of proteasomal degradation because it was sensitive to MG-132. The late reduction of Ha-Ras levels was abrogated by compound PD98059, which inhibits the MAP kinase pathway, whereas the late reduction of Ki-Ras was unaffected by PD98059. We conclude that oxidative stress differentially regulates the expression of Ha-Ras and Ki-Ras in cultured astrocytes, and that activation of the MAP kinase pathway by oxidative stress itself or by additional factors may act as a fail-safe mechanism limiting a sustained expression of the potentially detrimental Ha-Ras.

Amplification of oncogenes and integrated SV40 sequences in mammalian cells by the decay of incorporated iodine-125

International Nuclear Information System (INIS)

Ehrfeld, A.; Planas-Bohne, F.; Luecke-Huhle, C.

1986-01-01

Iodine-125, in the form of 5-[ 125 I]iododeoxyuridine (I-UdR), was incorporated into the DNA of SV40 transformed Chinese hamster embryo cells. Disintegration of the 125 I led to increased cell killing with increasing dose as measured by the colony-forming ability of single cells. The D37 (the dose at which 37% of the cells survive) amounts to 95 decays per cell, corresponding to 0.66 Gy. Variations in the copy number of specific DNA sequences was measured by using dispersed cell blotting with sensitive DNA hybridizations. A 13-fold amplification of the viral DNA sequences (SV40) and a twofold amplification of two cellular oncogenes of the ras-family (Ki-ras and Ha-ras) were found. Other cellular genes, like the alpha-actin gene, were not amplified, and no variation in gene copy number was detected after incubation of cells with cold I-UdR. We suggest the observed gene amplifications are induced by the densely ionizing radiation emitted by the decay of the incorporated 125 I atoms

p53, erbB-2 and K-ras gene alterations are rare in spontaneous and plutonium-239-induced canine lung neoplasia

International Nuclear Information System (INIS)

Tierney, L.A.; Hahn, F.F.; Lechner, J.F.

1996-01-01

Inhalation of high-linear energy transfer radiation in the form of radon progeny is a suspected cause of human lung cancer. To gain insight into the types of genetic derangements caused by this type of radiation, lung tumors from beagle dogs exposed to 239 PuO 2 and those arising in animals with no known carcinogen exposure were examined for evidence of aberrations in genes known to be altered in lung tumors. Altered expression of the p53 tumor suppressor gene and proto-oncogene erbB-2 proteins (p185 erbB2 ) was evaluated by immunohistochemical analysis of 117 tumors representing different histological types in exposed (n = 80) and unexposed (n = 37) animals. Twenty-eight tumors were analyzed for K-ras proto-oncogene mutations by polymerase chain reaction amplification and direct sequencing. Fourteen percent (16/116) of all lung neoplasms showed elevated nuclear accumulation of p53 protein. Regardless of exposure history, adenosquamous and squamous cell cancers comprised 94% of all tumors with p53 abnormalities. Eighteen percent (21/117) of all tumors had evidence of erbB-2 protein overexpression. K-ras mutations were not detected in codons 12, 13 or 61 of tumors from unexposed (n = 9) or plutonium-exposed dogs (n = 19). These data indicate that p53 and K-ras gene abnormalities as a result of missense mutation are infrequent events in spontaneous and 239 PuO 2 -induced lung neoplasia in this colony of beagle dogs. Alternative mechanisms of gene alteration may be involved in canine pulmonary carcinogenesis. 45 refs., 3 figs., 2 tabs

Malignant transformation of diploid human fibroblasts by transfection of oncogenes. Part 2, Progress report, July 1989--June 1992

Energy Technology Data Exchange (ETDEWEB)

McCormick, J.J.

1992-12-31

This document consist of brief reports prepared by postdoctoral students supported by the project, each describing his accomplishments under the grant. Topics include (1) Malignant Transformation of MSU-1. 1 Cells by Gamma Radiation, (2) Correlation between Levels of ras Expression and Presence of Transformed Phenotypes Including Tumorigenicity, Using a Modulatable Promoter, (3) Relation between Specific rad Oncogene Expression, (4) Correlation of Genetic Changes in Fibroblastic Tumors with Malignancies, (5)Transformation of MSU-1.1 Cells by sis Oncogene, (6) Malignant Transformation of MSU-1.0 Cells, (7) Correlation of Urokinase Plasminogen Activation (mu-PA) with Malignant Phenotype, (8)Two Dimensional Gel Electrophoresis Studies of the Proteins of the Major Cell Strains of the MSU-1 Family of Cells, and (9) Correlation between Proteinase Activity Levels and Malignancy.

Ras-Induced and Extracellular Signal-Regulated Kinase 1 and 2 Phosphorylation-Dependent Isomerization of Protein Tyrosine Phosphatase (PTP)-PEST by PIN1 Promotes FAK Dephosphorylation by PTP-PEST ▿

Science.gov (United States)

Zheng, Yanhua; Yang, Weiwei; Xia, Yan; Hawke, David; Liu, David X.; Lu, Zhimin

2011-01-01

Protein tyrosine phosphatase (PTP)-PEST is a critical regulator of cell adhesion and migration. However, the mechanism by which PTP-PEST is regulated in response to oncogenic signaling to dephosphorylate its substrates remains unclear. Here, we demonstrate that activated Ras induces extracellular signal-regulated kinase 1 and 2-dependent phosphorylation of PTP-PEST at S571, which recruits PIN1 to bind to PTP-PEST. Isomerization of the phosphorylated PTP-PEST by PIN1 increases the interaction between PTP-PEST and FAK, which leads to the dephosphorylation of FAK Y397 and the promotion of migration, invasion, and metastasis of v-H-Ras-transformed cells. These findings uncover an important mechanism for the regulation of PTP-PEST in activated Ras-induced tumor progression. PMID:21876001

A novel oncogenic BTK isoform is overexpressed in colon cancers and required for RAS-mediated transformation

DEFF Research Database (Denmark)

Grassilli, E; Pisano, F; Cialdella, A

2016-01-01

Bruton's tyrosine kinase (BTK) is essential for B-cell proliferation/differentiation and it is generally believed that its expression and function are limited to bone marrow-derived cells. Here, we report the identification and characterization of p65BTK, a novel isoform abundantly expressed in c...... therapeutic approach.Oncogene advance online publication, 25 January 2016; doi:10.1038/onc.2015.504....

RAS Insight

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David Heimbrook, now CEO of the Frederick National Laboratory for Cancer Research, played a major role in a large pharma as it tried to develop an anti-RAS drug. Lessons from that failure inform the RAS Initiative today.

A functional screen reveals an extensive layer of transcriptional and splicing control underlying RAS/MAPK signaling in Drosophila.

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Dariel Ashton-Beaucage

2014-03-01

Full Text Available The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway--including a new protein complex modulating RAF activation--we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing

Regulation of Ras exchange factors and cellular localization of Ras activation by lipid messengers in T cells

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Jesse E. Jun

2013-09-01

Full Text Available The Ras-MAPK signaling pathway is highly conserved throughout evolution and is activated downstream of a wide range of receptor stimuli. Ras guanine nucleotide exchange factors (RasGEFs catalyze GTP loading of Ras and play a pivotal role in regulating receptor-ligand induced Ras activity. In T cells, three families of functionally important RasGEFs are expressed: RasGRF, RasGRP, and SOS-family GEFs.Early on it was recognized that Ras activation is critical for T cell development and that the RasGEFs play an important role herein. More recent work has revealed that nuances in Ras activation appear to significantly impact T cell development and selection. These nuances include distinct biochemical patterns of analog versus digital Ras activation, differences in cellular localization of Ras activation, and intricate interplays between the RasGEFs during distinct T cell developmental stages as revealed by various new mouse models. In many instances, the exact nature of these nuances in Ras activation or how these may result from fine-tuning of the RasGEFs is not understood.One large group of biomolecules critically involved in the control of Ras-GEFs´functions are lipid second messengers. Multiple, yet distinct lipid products are generated following T cell receptor (TCR stimulation and bind to different domains in the RasGRP and SOS RasGEFs to facilitate the activation of the membrane-anchored Ras GTPases. In this review we highlight how different lipid-based elements are generated by various enzymes downstream of the TCR and other receptors and how these dynamic and interrelated lipid products may fine-tune Ras activation by RasGEFs in developing T cells.

EMT-induced stemness and tumorigenicity are fueled by the EGFR/Ras pathway.

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Dominic Chih-Cheng Voon

Full Text Available Recent studies have revealed that differentiated epithelial cells would acquire stem cell-like and tumorigenic properties following an Epithelial-Mesenchymal Transition (EMT. However, the signaling pathways that participate in this novel mechanism of tumorigenesis have not been fully characterized. In Runx3 (-/- p53 (-/- murine gastric epithelial (GIF-14 cells, EMT-induced plasticity is reflected in the expression of the embryonal proto-oncogene Hmga2 and Lgr5, an exclusive gastrointestinal stem cell marker. Here, we report the concurrent activation of an EGFR/Ras gene expression signature during TGF-β1-induced EMT in GIF-14 cells. Amongst the altered genes was the induction of Egfr, which corresponded with a delayed sensitization to EGF treatment in GIF-14. Co-treatment with TGF-β1 and EGF or the expression of exogenous KRas led to increased Hmga2 or Lgr5 expression, sphere initiation and colony formation in soft agar assay. Interestingly, the gain in cellular plasticity/tumorigenicity was not accompanied by increased EMT. This uncoupling of EMT and the induction of plasticity reveals an involvement of distinct signaling cues, whereby the EGFR/Ras pathway specifically promotes stemness and tumorigenicity in EMT-altered GIF-14 cells. These data show that the EGFR/Ras pathway requisite for the sustenance of gastric stem cells in vivo and in vitro is involved in the genesis and promotion of EMT-induced tumor-initiating cells.

Antisense gene therapy using anti-k-ras and antitelomerase oligonucleotides in colorectal cancer Eficacia de la terapia génica antisentido utilizando oligonucleótidos anti K-ras y antitelomerasa en cáncer colorrectal

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S. Lledó

2005-07-01

Full Text Available Aim: to test the efficacy of anti-k-ras and antitelomerase oligonucleotides for disabling colorectal cancer cell growth. Material and methods: an established human colorectal cancer cell line (SW 480, ATTC® was used. Oligodeoxiribonucleotides (ODNs have a phosphorotioate modification to ensure intracellular intake. We used an antitelomerase ODN (Telp5 and two anti-k-ras ODNs (AS-KRAS and ISIS. AS-KRAS is designed to join the k-ras oncogene's exon 1. ISIS links to the terminal transcription unit 5' of k-ras. Telp5 joins the template region of the hTR telomerase subunit. ODNs have been tested in different concentrations (1, 5, 10, 20 micromolar. Cell viability has been tested at 48 and 72 hours. Statistical analysis and graphic design were made with the statistical package "Analyzing Data with GraphPad Prism-1999", GraphPad Sofware Inc., San Diego CA©. We used the Student's t test for statistical analysis. Results: the lowest dose (1 µM was not effective. Using the highest dose (20 mM for 48 hours of combined AS-KRAS and Telp5 cell viability decreased to 99.67%. The rest of results varied depending on ODN type, dose, and exposure time. Conclusions: tested antisense ODNs stop colorectal cancer cell growth, and a combination of anti-telomerase and anti-k-ras is the most useful treatment. Efficacy is best with a higher dose and longer treatment period.Objetivo: evaluar la eficacia de oligonucleótidos anti k-ras y antitelomerasa para detener el crecimiento tumoral en el cáncer colorrectal. Material y métodos: se ha empleado una línea celular establecida de cáncer colorrectal humano (SW 480, ATTC®. Los oligodesoxirribonucleótidos (ODN utilizados en el presente trabajo presentan modificación fosforotioato con el fin de mejorar su estabilidad en presencia de fluidos biológicos. Hemos utilizado un ODN antitelomerasa (Telp5, y dos ODN anti k-ras (AS-KRAS e ISIS. AS-KRAS actúa en el exón 1 e ISIS actúa a nivel de la unidad terminal de

RAS - Target Identification - Informatics

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The RAS Informatics lab group develops tools to track and analyze “big data” from the RAS Initiative, as well as analyzes data from external projects. By integrating internal and external data, this group helps improve understanding of RAS-driven cancers.

Identification of an intracellular protein that specifically interacts with photoaffinity-labeled oncogenic p21 protein

International Nuclear Information System (INIS)

Lee, G.; Ronai, Z.A.; Pincus, M.R.; Brandt-Rauf, P.W.; Weinstein, I.B.; Murphy, R.B.; Delohery, T.M.; Nishimura, S.; Yamaizumi, Z.

1989-01-01

An oncogenic 21-kDa (p21) protein (Harvey RAS protein with Val-12) has been covalently modified with a functional reagent that contains a photoactivatable aromatic azide group. This modified p21 protein has been introduced quantitatively into NIH 3T3 cells using an erythrocyte-mediated fusion technique. The introduced p21 protein was capable of inducing enhanced pinocytosis and DNA synthesis in the recipient cells. To identify the putative intracellular protein(s) that specifically interact with modified p21 protein, the cells were pulsed with [ 35 S]methionine at selected times after fusion and then UV-irradiated to activate the azide group. The resulting nitrene covalently binds to amino acid residues in adjacent proteins, thus linking the p21 protein to these proteins. The cells were then lysed, and the lysate was immunoprecipitated with the anti-p21 monoclonal antibody Y13-259. The immunoprecipitate was analyzed by SDS/PAGE to identify p21 - protein complexes. By using this technique, the authors found that three protein complexes of 51, 64, and 82 kDa were labeled specifically and reproducibly. The most prominent band is the 64-kDa protein complex that shows a time-dependent rise and fall, peaking within a 5-hr period after introduction of the p21 protein the cells. These studies provide evidence that in vitro the p21 protein becomes associated with a protein whose mass is about 43 kDa. They suggest that the formation of this complex may play a role in mediating early events involved with cell transformation induced by RAS oncogenes

Inhibitors of Ras-SOS Interactions.

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Lu, Shaoyong; Jang, Hyunbum; Zhang, Jian; Nussinov, Ruth

2016-04-19

Activating Ras mutations are found in about 30 % of human cancers. Ras activation is regulated by guanine nucleotide exchange factors, such as the son of sevenless (SOS), which form protein-protein interactions (PPIs) with Ras and catalyze the exchange of GDP by GTP. This is the rate-limiting step in Ras activation. However, Ras surfaces lack any evident suitable pockets where a molecule might bind tightly, rendering Ras proteins still 'undruggable' for over 30 years. Among the alternative approaches is the design of inhibitors that target the Ras-SOS PPI interface, a strategy that is gaining increasing recognition for treating Ras mutant cancers. Herein we focus on data that has accumulated over the past few years pertaining to the design of small-molecule modulators or peptide mimetics aimed at the interface of the Ras-SOS PPI. We emphasize, however, that even if such Ras-SOS therapeutics are potent, drug resistance may emerge. To counteract this development, we propose "pathway drug cocktails", that is, drug combinations aimed at parallel (or compensatory) pathways. A repertoire of classified cancer, cell/tissue, and pathway/protein combinations would be beneficial toward this goal. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Small GTPase R-Ras participates in neural tube formation in zebrafish embryonic spinal cord.

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Ohata, Shinya; Uga, Hideko; Okamoto, Hitoshi; Katada, Toshiaki

2018-06-27

Ras related (R-Ras), a small GTPase, is involved in the maintenance of apico-basal polarity in neuroepithelial cells of the zebrafish hindbrain, axonal collapse in cultured murine hippocampal neurons, and maturation of blood vessels in adult mice. However, the role of R-Ras in neural tube formation remains unknown. Using antisense morpholino oligonucleotides (AMOs), we found that in the spinal cord of zebrafish embryos, the lumen was formed bilaterally in rras morphants, whereas it was formed at the midline in control embryos. As AMO can cause off-target effects, we generated rras mutant zebrafish lines using CRISPR/Cas9 technology. Although these rras mutant embryos did not have a bilateral lumen in the spinal cord, the following findings suggest that the phenotype is unlikely due to an off-target effect of rras AMO: 1) The rras morphant phenotype was rescued by an injection of AMO-resistant rras mRNA, and 2) a bilaterally segregated spinal cord was not observed in rras mutant embryos injected with rras AMO. The results suggest that the function of other ras family genes may be redundant in rras mutants. Previous research reported a bilaterally formed lumen in the spinal cord of zebrafish embryos with a mutation in a planar cell polarity (PCP) gene, van gogh-like 2 (vangl2). In the present study, in cultured cells, R-Ras was co-immunoprecipitated with Vangl2 but not with another PCP regulator, Pricke1. Interestingly, the interaction between R-Ras and Vangl2 was stronger in guanine-nucleotide free point mutants of R-Ras than in wild-type or constitutively active (GTP-bound) forms of R-Ras. R-Ras may regulate neural tube formation in cooperation with Vangl2 in the developing zebrafish spinal cord. Copyright © 2018 Elsevier Inc. All rights reserved.

Oncogenes, radiation and cancer; Oncogenes, radiacion y cancer

Energy Technology Data Exchange (ETDEWEB)

Michelin, S C

1999-12-31

The discovery of the oncogenic virus and the analysis of its nucleic acid, together with the development of new biochemical technology have permitted the partial knowledge of the molecular mechanisms responsible for the cellular neoplastic transformation. This work, besides describing the discovery of the first oncogenic virus and the experiments to demonstrate the existence of the oncogenes, summarizes its activation mechanisms and its intervention in cellular metabolisms. Ionizing radiation is among the external agents that induce the neoplastic process. Its participation in the genesis of this process and the contribution of oncogenes to the cellular radioresistance are among the topics, which are referred to another topic that makes reference. At the same time as the advancement of theoretical knowledge, lines of investigation for the application of the new concepts in diagnosis, prognosis and therapeutical treatment, were developed. An example of this, is the study of the participation of the oncogen c-erbB-2 in human breast cancer and its implications on the anti tumoral therapy. (author) 87 refs., 7 figs., 3 tabs. [Espanol] El descubrimiento de los virus oncogenicos y el analisis de su acido nucleico, junto con el desarrollo de nuevas tecnicas bioquimicas, ha permitido conocer parcialmente los mecanismos moleculares responsables de la transformacion de una celula normal en neoplasica. En este trabajo, ademas de describir el descubrimiento de los primeros virus oncogenicos y las experiencias para demostrar la existencia de los oncogenes, se resumen sus mecanismos de activacion y su intervencion en el metabolismo celular. Entre los agentes expernos que inducen un proceso oncogenico, se encuentran las radiaciones ionizantes. Su participacion en la genesis de este proceso y la contribucion de los oncogenes a la radioresistencia de las celulas tumorales, es otro de los temas a que se hace referencia. Paralelamente al avance del conocimiento teorico, se

RAS Initiative - Events

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The NCI RAS Initiative has organized multiple events with outside experts to discuss how the latest scientific and technological breakthroughs can be applied to discover vulnerabilities in RAS-driven cancers.

Cyclic AMP signalling in Dictyostelium : G-proteins activate separate Ras pathways using specific RasGEFs

NARCIS (Netherlands)

Kae, Helmut; Kortholt, Arjan; Rehmann, Holger; Insall, RobertH.; Van Haastert, Peter J. M.; Spiegelman, George B.; Weeks, Gerald

In general, mammalian Ras guanine nucleotide exchange factors (RasGEFs) show little substrate specificity, although they are often thought to regulate specific pathways. Here, we provide in vitro and in vivo evidence that two RasGEFs can each act on specific Ras proteins. During Dictyostelium

Transforming p21 ras protein: flexibility in the major variable region linking the catalytic and membrane-anchoring domains

DEFF Research Database (Denmark)

Willumsen, B M; Papageorge, A G; Hubbert, N

1985-01-01

or increasing it to 50 amino acids has relatively little effect on the capacity of the gene to induce morphological transformation of NIH 3T3 cells. Assays of GTP binding, GTPase and autophosphorylating activities of such mutant v-rasH-encoded proteins synthesized in bacteria indicated that the sequences...... that is required for post-translational processing, membrane localization and transforming activity of the proteins. We have now used the viral oncogene (v-rasH) of Harvey sarcoma virus to study the major variable region by deleting or duplicating parts of the gene. Reducing this region to five amino acids...... that encode these biochemical activities are located upstream from the major variable region. In the context of transformation, we propose that the region of sequence heterogeneity serves principally to connect the N-terminal catalytic domain with amino acids at the C terminus that are required to anchor...

A gene expression signature of RAS pathway dependence predicts response to PI3K and RAS pathway inhibitors and expands the population of RAS pathway activated tumors.

Science.gov (United States)

Loboda, Andrey; Nebozhyn, Michael; Klinghoffer, Rich; Frazier, Jason; Chastain, Michael; Arthur, William; Roberts, Brian; Zhang, Theresa; Chenard, Melissa; Haines, Brian; Andersen, Jannik; Nagashima, Kumiko; Paweletz, Cloud; Lynch, Bethany; Feldman, Igor; Dai, Hongyue; Huang, Pearl; Watters, James

2010-06-30

Hyperactivation of the Ras signaling pathway is a driver of many cancers, and RAS pathway activation can predict response to targeted therapies. Therefore, optimal methods for measuring Ras pathway activation are critical. The main focus of our work was to develop a gene expression signature that is predictive of RAS pathway dependence. We used the coherent expression of RAS pathway-related genes across multiple datasets to derive a RAS pathway gene expression signature and generate RAS pathway activation scores in pre-clinical cancer models and human tumors. We then related this signature to KRAS mutation status and drug response data in pre-clinical and clinical datasets. The RAS signature score is predictive of KRAS mutation status in lung tumors and cell lines with high (> 90%) sensitivity but relatively low (50%) specificity due to samples that have apparent RAS pathway activation in the absence of a KRAS mutation. In lung and breast cancer cell line panels, the RAS pathway signature score correlates with pMEK and pERK expression, and predicts resistance to AKT inhibition and sensitivity to MEK inhibition within both KRAS mutant and KRAS wild-type groups. The RAS pathway signature is upregulated in breast cancer cell lines that have acquired resistance to AKT inhibition, and is downregulated by inhibition of MEK. In lung cancer cell lines knockdown of KRAS using siRNA demonstrates that the RAS pathway signature is a better measure of dependence on RAS compared to KRAS mutation status. In human tumors, the RAS pathway signature is elevated in ER negative breast tumors and lung adenocarcinomas, and predicts resistance to cetuximab in metastatic colorectal cancer. These data demonstrate that the RAS pathway signature is superior to KRAS mutation status for the prediction of dependence on RAS signaling, can predict response to PI3K and RAS pathway inhibitors, and is likely to have the most clinical utility in lung and breast tumors.

A gene expression signature of RAS pathway dependence predicts response to PI3K and RAS pathway inhibitors and expands the population of RAS pathway activated tumors

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Paweletz Cloud

2010-06-01

Full Text Available Abstract Background Hyperactivation of the Ras signaling pathway is a driver of many cancers, and RAS pathway activation can predict response to targeted therapies. Therefore, optimal methods for measuring Ras pathway activation are critical. The main focus of our work was to develop a gene expression signature that is predictive of RAS pathway dependence. Methods We used the coherent expression of RAS pathway-related genes across multiple datasets to derive a RAS pathway gene expression signature and generate RAS pathway activation scores in pre-clinical cancer models and human tumors. We then related this signature to KRAS mutation status and drug response data in pre-clinical and clinical datasets. Results The RAS signature score is predictive of KRAS mutation status in lung tumors and cell lines with high (> 90% sensitivity but relatively low (50% specificity due to samples that have apparent RAS pathway activation in the absence of a KRAS mutation. In lung and breast cancer cell line panels, the RAS pathway signature score correlates with pMEK and pERK expression, and predicts resistance to AKT inhibition and sensitivity to MEK inhibition within both KRAS mutant and KRAS wild-type groups. The RAS pathway signature is upregulated in breast cancer cell lines that have acquired resistance to AKT inhibition, and is downregulated by inhibition of MEK. In lung cancer cell lines knockdown of KRAS using siRNA demonstrates that the RAS pathway signature is a better measure of dependence on RAS compared to KRAS mutation status. In human tumors, the RAS pathway signature is elevated in ER negative breast tumors and lung adenocarcinomas, and predicts resistance to cetuximab in metastatic colorectal cancer. Conclusions These data demonstrate that the RAS pathway signature is superior to KRAS mutation status for the prediction of dependence on RAS signaling, can predict response to PI3K and RAS pathway inhibitors, and is likely to have the most clinical

Kita driven expression of oncogenic HRAS leads to early onset and highly penetrant melanoma in zebrafish.

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Cristina Santoriello

2010-12-01

Full Text Available Melanoma is the most aggressive and lethal form of skin cancer. Because of the increasing incidence and high lethality of melanoma, animal models for continuously observing melanoma formation and progression as well as for testing pharmacological agents are needed.Using the combinatorial Gal4-UAS system, we have developed a zebrafish transgenic line that expresses oncogenic HRAS under the kita promoter. Already at 3 days transgenic kita-GFP-RAS larvae show a hyper-pigmentation phenotype as earliest evidence of abnormal melanocyte growth. By 2-4 weeks, masses of transformed melanocytes form in the tail stalk of the majority of kita-GFP-RAS transgenic fish. The adult tumors evident between 1-3 months of age faithfully reproduce the immunological, histological and molecular phenotypes of human melanoma, but on a condensed time-line. Furthermore, they show transplantability, dependence on mitfa expression and do not require additional mutations in tumor suppressors. In contrast to kita expressing melanocyte progenitors that efficiently develop melanoma, mitfa expressing progenitors in a second Gal4-driver line were 4 times less efficient in developing melanoma during the three months observation period.This indicates that zebrafish kita promoter is a powerful tool for driving oncogene expression in the right cells and at the right level to induce early onset melanoma in the presence of tumor suppressors. Thus our zebrafish model provides a link between kita expressing melanocyte progenitors and melanoma and offers the advantage of a larval phenotype suitable for large scale drug and genetic modifier screens.

Genetic alterations in Ki-ras and Ha-ras genes in Juvenile Nasopharyngeal Angiofibromas and head and neck cancer

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Cláudia Malheiros Coutinho

1999-05-01

Full Text Available CONETXT: Ras gene mutations have been associated to a wide range of human solid tumors. Members of the ras gene family (Ki-ras, Ha-ras and N-ras are structurally related and code for a protein (p21 known to play an important role in the regulation of normal signal transduction and cell growth. The frequency of ras mutations is different from one type of tumor to another, suggesting that point mutations might be carcinogen-specific. OBJECTIVES: To study the occurrence of Ki-ras and Ha-ras mutations. We also studied the relative level of Ha-ras mRNA in 32 of the head and neck tumors. DESIGN: Case series. SETTING: University referral unit. PARTICIPANTS: 60 head and neck tumors and in 28 Juvenile Nasopharyngeal Angiofibromas (JNA. DIAGNOSTIC TEST: Using PCR-SSCP we examined the occurrence of Ki-ras and Ha-ras mutations. The relative level of Ha-ras mRNA was examined by Northern blot analysis. RESULTS: None of the head and neck tumors or JNA samples showed evidence of mutations within codons 12, 13, 59 and 61 of Ki-ras or Ha-ras genes. However, 17 (53% of the tumors where gene expression could be examined exhibited increased levels of Ha-ras mRNA compared with the normal tissue derived from the same patient. CONCLUSIONS: Our results demonstrate for the first time that mutations of Ki-ras and Ha-ras genes are not associated with the development of JNA and confirm previous reports indicating that activating ras mutations are absent or rarely involved in head and neck tumors from western world patients. Furthermore, our findings suggest that overexpression of Ha-ras, rather than mutations, might be an important factor in the development and progression of head and neck tumors.

The oncogenic RAS2 val19 mutation locks respiration, independently of PKA, in a mode prone to generate ROS

Czech Academy of Sciences Publication Activity Database

Hlavatá, Lydie; Aguilaniu, H.; Pichová, Alena; Nystrom, T.

2003-01-01

Roč. 22, č. 13 (2003), s. 3337-3345 ISSN 0261-4189 R&D Projects: GA ČR GA301/03/0289 Institutional research plan: CEZ:AV0Z5020903 Keywords : pka/ras2/ * ucp1 * respiratory state Subject RIV: EE - Microbiology, Virology Impact factor: 10.456, year: 2003

Fetisisme Ras Kaukasoid dan Ras Mongoloid Sebagai Strategi Pemasaran dalam Sinetron Indonesia

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Dimas Yudhistira

2014-12-01

ABSTRAK   Budaya populer yang tumbuh seiring dengan industrialisasi memengaruhi produksi per- filman di Indonesia. Salah satu genre perfilman di Indonesia adalah sinetron. Sinetron yang di- kategorikan sebagai produk seni kitsch memiliki dua kriteria yaitu sebagai komoditi seni yang populer dan sebagai komoditi dagang yang menghasilkan keuntungan ekonomis. Sebagai se- buah produk seni kitsch yang merupakan dasar pembuatan karyanya adalah selera masyarakat kebanyakan maka sinetron harus jeli dalam melihat keadaan dan latar belakang masyarakat. Penelitian ini menggunakan metode kualitatif. Hasil penelitian ini menggambarkan masyara- kat Indonesia yang merupakan ras Melayu telah dijajah oleh ras Kaukasoid dan Mongoloid sebelum tahun 1945 dan setelahnya. Efek dari penjajahan ini adalah ras Melayu telah ditanami fantasi yang menjadi stereotip mengenai ras Kaukasoid dan Mongoloid yang berakhir dengan fetisisme. Fetisisme ini dijadikan sebagai strategi pemasaran oleh produser dan sutradara un- tuk menarik antusiasme calon penonton sinetron. Caranya dengan menampilkan aktor dan aktris Melayu keturunan Kaukasoid dan Mongoloid sebagai pemeran utama.   Kata kunci: sinetron, seni kitsch, ras, fetisisme

Regulation of p21ras activity

DEFF Research Database (Denmark)

Lowy, D R; Zhang, K; DeClue, J E

1992-01-01

The ras genes encode GTP/GDP-binding proteins that participate in mediating mitogenic signals from membrane tyrosine kinases to downstream targets. The activity of p21ras is determined by the concentration of GTP-p21ras, which is tightly regulated by a complex array of positive and negative control...... mechanisms. GAP and NF1 can negatively regulate p21ras activity by stimulating hydrolysis of GTP bound to p21ras. Other cellular factors can positively regulate p21ras by stimulating GDP/GTP exchange....

Identification of an intracellular protein that specifically interacts with photoaffinity-labeled oncogenic p21 protein.

Science.gov (United States)

Lee, G; Ronai, Z A; Pincus, M R; Brandt-Rauf, P W; Murphy, R B; Delohery, T M; Nishimura, S; Yamaizumi, Z; Weinstein, I B

1989-11-01

An oncogenic 21-kDa (p21) protein (Harvey RAS protein with Val-12) has been covalently modified with a functional reagent that contains a photoactivatable aromatic azide group. This modified p21 protein has been introduced quantitatively into NIH 3T3 cells using an erythrocyte-mediated fusion technique. The introduced p21 protein was capable of inducing enhanced pinocytosis and DNA synthesis in the recipient cells. To identify the putative intracellular protein(s) that specifically interact with the modified p21 protein, the cells were pulsed with [35S]methionine at selected times after fusion and then UV-irradiated to activate the azide group. The resulting nitrene covalently binds to amino acid residues in adjacent proteins, thus linking the p21 protein to these proteins. The cells were then lysed, and the lysate was immunoprecipitated with the anti-p21 monoclonal antibody Y13-259. The immunoprecipitate was analyzed by SDS/PAGE to identify p21-protein complexes. By using this technique, we found that three protein complexes of 51, 64, and 82 kDa were labeled specifically and reproducibly. The most prominent band is the 64-kDa protein complex that shows a time-dependent rise and fall, peaking within a 5-hr period after introduction of the p21 protein into the cells. These studies provide evidence that in vitro the p21 protein becomes associated with a protein whose mass is about 43 kDa. We suggest that the formation of this complex may play a role in mediating early events involved with cell transformation induced by RAS oncogenes.

A Histidine pH sensor regulates activation of the Ras-specific guanine nucleotide exchange factor RasGRP1.

Science.gov (United States)

Vercoulen, Yvonne; Kondo, Yasushi; Iwig, Jeffrey S; Janssen, Axel B; White, Katharine A; Amini, Mojtaba; Barber, Diane L; Kuriyan, John; Roose, Jeroen P

2017-09-27

RasGRPs are guanine nucleotide exchange factors that are specific for Ras or Rap, and are important regulators of cellular signaling. Aberrant expression or mutation of RasGRPs results in disease. An analysis of RasGRP1 SNP variants led to the conclusion that the charge of His 212 in RasGRP1 alters signaling activity and plasma membrane recruitment, indicating that His 212 is a pH sensor that alters the balance between the inactive and active forms of RasGRP1. To understand the structural basis for this effect we compared the structure of autoinhibited RasGRP1, determined previously, to those of active RasGRP4:H-Ras and RasGRP2:Rap1b complexes. The transition from the autoinhibited to the active form of RasGRP1 involves the rearrangement of an inter-domain linker that displaces inhibitory inter-domain interactions. His 212 is located at the fulcrum of these conformational changes, and structural features in its vicinity are consistent with its function as a pH-dependent switch.

Mapping the Conformation Space of Wildtype and Mutant H-Ras with a Memetic, Cellular, and Multiscale Evolutionary Algorithm.

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Rudy Clausen

2015-09-01

Full Text Available An important goal in molecular biology is to understand functional changes upon single-point mutations in proteins. Doing so through a detailed characterization of structure spaces and underlying energy landscapes is desirable but continues to challenge methods based on Molecular Dynamics. In this paper we propose a novel algorithm, SIfTER, which is based instead on stochastic optimization to circumvent the computational challenge of exploring the breadth of a protein's structure space. SIfTER is a data-driven evolutionary algorithm, leveraging experimentally-available structures of wildtype and variant sequences of a protein to define a reduced search space from where to efficiently draw samples corresponding to novel structures not directly observed in the wet laboratory. The main advantage of SIfTER is its ability to rapidly generate conformational ensembles, thus allowing mapping and juxtaposing landscapes of variant sequences and relating observed differences to functional changes. We apply SIfTER to variant sequences of the H-Ras catalytic domain, due to the prominent role of the Ras protein in signaling pathways that control cell proliferation, its well-studied conformational switching, and abundance of documented mutations in several human tumors. Many Ras mutations are oncogenic, but detailed energy landscapes have not been reported until now. Analysis of SIfTER-computed energy landscapes for the wildtype and two oncogenic variants, G12V and Q61L, suggests that these mutations cause constitutive activation through two different mechanisms. G12V directly affects binding specificity while leaving the energy landscape largely unchanged, whereas Q61L has pronounced, starker effects on the landscape. An implementation of SIfTER is made available at http://www.cs.gmu.edu/~ashehu/?q=OurTools. We believe SIfTER is useful to the community to answer the question of how sequence mutations affect the function of a protein, when there is an

About the RAS Initiative

Science.gov (United States)

The RAS Initiative, a "hub and spoke" model, connects researchers to better understand and target the more than 30% of cancers driven by mutations in RAS genes. Includes oversight and contact information.

Inactivation of the DNA repair gene O6-methylguanine-DNA methyltransferase by promoter hypermethylation is associated with G to A mutations in K-ras in colorectal tumorigenesis.

Science.gov (United States)

Esteller, M; Toyota, M; Sanchez-Cespedes, M; Capella, G; Peinado, M A; Watkins, D N; Issa, J P; Sidransky, D; Baylin, S B; Herman, J G

2000-05-01

O6-methylguanine DNA methyltransferase (MGMT) is a DNA repair protein that removes mutagenic and cytotoxic adducts from the O6 position of guanine. O6-methylguanine mispairs with thymine during replication, and if the adduct is not removed, this results in conversion from a guanine-cytosine pair to an adenine-thymine pair. In vitro assays show that MGMT expression avoids G to A mutations and MGMT transgenic mice are protected against G to A transitions at ras genes. We have recently demonstrated that the MGMT gene is silenced by promoter methylation in many human tumors, including colorectal carcinomas. To study the relevance of defective MGMT function by aberrant methylation in relation to the presence of K-ras mutations, we studied 244 colorectal tumor samples for MGMT promoter hypermethylation and K-ras mutational status. Our results show a clear association between the inactivation of MGMT by promoter hypermethylation and the appearance of G to A mutations at K-ras: 71% (36 of 51) of the tumors displaying this particular type of mutation had abnormal MGMT methylation, whereas only 32% (12 of 37) of those with other K-ras mutations not involving G to A transitions and 35% (55 of 156) of the tumors without K-ras mutations demonstrated MGMT methylation (P = 0.002). In addition, MGMT loss associated with hypermethylation was observed in the small adenomas, including those that do not yet contain K-ras mutations. Hypermethylation of other genes such as p16INK4a and p14ARF was not associated with either MGMT hypermethylation or K-ras mutation. Our data suggest that epigenetic silencing of MGMT by promoter hypermethylation may lead to a particular genetic change in human cancer, specifically G to A transitions in the K-ras oncogene.

What makes Ras an efficient molecular switch: a computational, biophysical, and structural study of Ras-GDP interactions with mutants of Raf.

Science.gov (United States)

Filchtinski, Daniel; Sharabi, Oz; Rüppel, Alma; Vetter, Ingrid R; Herrmann, Christian; Shifman, Julia M

2010-06-11

Ras is a small GTP-binding protein that is an essential molecular switch for a wide variety of signaling pathways including the control of cell proliferation, cell cycle progression and apoptosis. In the GTP-bound state, Ras can interact with its effectors, triggering various signaling cascades in the cell. In the GDP-bound state, Ras looses its ability to bind to known effectors. The interaction of the GTP-bound Ras (Ras(GTP)) with its effectors has been studied intensively. However, very little is known about the much weaker interaction between the GDP-bound Ras (Ras(GDP)) and Ras effectors. We investigated the factors underlying the nucleotide-dependent differences in Ras interactions with one of its effectors, Raf kinase. Using computational protein design, we generated mutants of the Ras-binding domain of Raf kinase (Raf) that stabilize the complex with Ras(GDP). Most of our designed mutations narrow the gap between the affinity of Raf for Ras(GTP) and Ras(GDP), producing the desired shift in binding specificity towards Ras(GDP). A combination of our best designed mutation, N71R, with another mutation, A85K, yielded a Raf mutant with a 100-fold improvement in affinity towards Ras(GDP). The Raf A85K and Raf N71R/A85K mutants were used to obtain the first high-resolution structures of Ras(GDP) bound to its effector. Surprisingly, these structures reveal that the loop on Ras previously termed the switch I region in the Ras(GDP).Raf mutant complex is found in a conformation similar to that of Ras(GTP) and not Ras(GDP). Moreover, the structures indicate an increased mobility of the switch I region. This greater flexibility compared to the same loop in Ras(GTP) is likely to explain the natural low affinity of Raf and other Ras effectors to Ras(GDP). Our findings demonstrate that an accurate balance between a rigid, high-affinity conformation and conformational flexibility is required to create an efficient and stringent molecular switch. Copyright 2010 Elsevier Ltd

Analysis of the K-ras and p53 pathways in x-ray-induced lung tumors in the rat

Energy Technology Data Exchange (ETDEWEB)

Belinsky, S.A.; Middleton, S.K.; Hahn, F.F.; Nikula, K.J. [Inhalation Toxicology Research Inst., Albuquerque, NM (United States); Picksley, S.M. [Medical Sciences Inst., Dundee (United Kingdom)

1996-04-01

The risk from exposure to low-dose radiation in conjunction with cigarette smoking has not been estimated due in part to lmited knowledge surrounding the molecular mechanisms underlying radiation-induced cancers. The purpose of this investigation was to determine the frequency for alterations in genes within the K-ras and p53 signal and cell cycle regulatory pathways, respectively, in X-ray-induced lung tumors in the F344/N rat. These tumors were examined for genetic alterations in the K-ras, c-raf-1, p53, mdm2 and cip1 genes. No K-ras mutations were detected by sequencing in 18 squamous cell carcinomas (SCCs) or 17 adenocarcinomas. However, using a K-ras codon 12 mutation selection assay, a codon 12 GGT {r_arrow} GAT mutation was detected in one SCC, suggesting that activation of the K-ras proto-oncogene is both a rare and late event. Single-strand conformation polymorphism (SSCP) analysis of the kinase-binding domain of the c-raf-1 gene did not detect any polymorphisms. Three of 18 SCCs but none of the adenocarcinomas showed p53 nuclear immunoreactivity. Single-strand conformation polymorphism analysis of exons 4-9 of the p53 gene detected only an exon 9 mutation in one SCC. Mutations were not detected in the three SCCs with immunoreactive p53 protein. No amplification of the mdm2 gene was detected; however, nuclear mdm2 immunoreactivity was present in one of the three SCCs that stained positive for the p53 protein. The complete cDNA of the rat cip1 gene comprising 810 bases was cloned and sequenced. The frequency of somatic mutations in exon 2 of the cip1 gene was determined by SSCP analysis. No alterations in electrophoretic mobility were detected. The results of this investigation indicate that alterations in the K-ras and p53 pathways do not play a major role in the genesis of X-ray-induced lung tumors in the rat. 49 refs., 5 figs.

RAS Initiative - Community Outreach

Science.gov (United States)

Through community and technical collaborations, workshops and symposia, and the distribution of reference reagents, the RAS Initiative seeks to increase the sharing of knowledge and resources essential to defeating cancers caused by mutant RAS genes.

The C-terminus of H-Ras as a target for the covalent binding of reactive compounds modulating Ras-dependent pathways.

Directory of Open Access Journals (Sweden)

Clara L Oeste

2011-01-01

Full Text Available Ras proteins are crucial players in differentiation and oncogenesis and constitute important drug targets. The localization and activity of Ras proteins are highly dependent on posttranslational modifications at their C-termini. In addition to an isoprenylated cysteine, H-Ras, but not other Ras proteins, possesses two cysteine residues (C181 and C184 in the C-terminal hypervariable domain that act as palmitoylation sites in cells. Cyclopentenone prostaglandins (cyPG are reactive lipidic mediators that covalently bind to H-Ras and activate H-Ras dependent pathways. Dienone cyPG, such as 15-deoxy-Δ(12,14-PGJ(2 (15d-PGJ(2 and Δ(12-PGJ(2 selectively bind to the H-Ras hypervariable domain. Here we show that these cyPG bind simultaneously C181 and C184 of H-Ras, thus potentially altering the conformational tendencies of the hypervariable domain. Based on these results, we have explored the capacity of several bifunctional cysteine reactive small molecules to bind to the hypervariable domain of H-Ras proteins. Interestingly, phenylarsine oxide (PAO, a widely used tyrosine phosphatase inhibitor, and dibromobimane, a cross-linking agent used for cysteine mapping, effectively bind H-Ras hypervariable domain. The interaction of PAO with H-Ras takes place in vitro and in cells and blocks modification of H-Ras by 15d-PGJ(2. Moreover, PAO treatment selectively alters H-Ras membrane partition and the pattern of H-Ras activation in cells, from the plasma membrane to endomembranes. These results identify H-Ras as a novel target for PAO. More importantly, these observations reveal that small molecules or reactive intermediates interacting with spatially vicinal cysteines induce intramolecular cross-linking of H-Ras C-terminus potentially contributing to the modulation of Ras-dependent pathways.

Oncogenic programmes and Notch activity: an 'organized crime'?

Science.gov (United States)

Dominguez, Maria

2014-04-01

The inappropriate Notch signalling can influence virtually all aspect of cancer, including tumour-cell growth, survival, apoptosis, angiogenesis, invasion and metastasis, although it does not do this alone. Hence, elucidating the partners of Notch that are active in cancer is now the focus of much intense research activity. The genetic toolkits available, coupled to the small size and short life of the fruit fly Drosophila melanogaster, makes this an inexpensive and effective animal model, suited to large-scale cancer gene discovery studies. The fly eye is not only a non-vital organ but its stereotyped size and disposition also means it is easy to screen for mutations that cause tumours and metastases and provides ample opportunities to test cancer theories and to unravel unanticipated nexus between Notch and other cancer genes, or to discover unforeseen Notch's partners in cancer. These studies suggest that Notch's oncogenic capacity is brought about not simply by increasing signal strength but through partnerships, whereby oncogenes gain more by cooperating than acting individually, as in a ring 'organized crime'. Copyright © 2014 Elsevier Ltd. All rights reserved.

Ras proteins have multiple functions in vegetative cells of Dictyostelium.

Science.gov (United States)

Bolourani, Parvin; Spiegelman, George; Weeks, Gerald

2010-11-01

During the aggregation of Dictyostelium cells, signaling through RasG is more important in regulating cyclic AMP (cAMP) chemotaxis, whereas signaling through RasC is more important in regulating the cAMP relay. However, RasC is capable of substituting for RasG for chemotaxis, since rasG⁻ cells are only partially deficient in chemotaxis, whereas rasC⁻/rasG⁻ cells are totally incapable of chemotaxis. In this study we have examined the possible functional overlap between RasG and RasC in vegetative cells by comparing the vegetative cell properties of rasG⁻, rasC⁻, and rasC⁻/rasG⁻ cells. In addition, since RasD, a protein not normally found in vegetative cells, is expressed in vegetative rasG⁻ and rasC⁻/rasG⁻ cells and appears to partially compensate for the absence of RasG, we have also examined the possible functional overlap between RasG and RasD by comparing the properties of rasG⁻ and rasC⁻/rasG⁻ cells with those of the mutant cells expressing higher levels of RasD. The results of these two lines of investigation show that RasD is capable of totally substituting for RasG for cytokinesis and growth in suspension, whereas RasC is without effect. In contrast, for chemotaxis to folate, RasC is capable of partially substituting for RasG, but RasD is totally without effect. Finally, neither RasC nor RasD is able to substitute for the role that RasG plays in regulating actin distribution and random motility. These specificity studies therefore delineate three distinct and none-overlapping functions for RasG in vegetative cells.

Rasputin, the Drosophila homologue of the RasGAP SH3 binding protein, functions in ras- and Rho-mediated signaling.

Science.gov (United States)

Pazman, C; Mayes, C A; Fanto, M; Haynes, S R; Mlodzik, M

2000-04-01

The small GTPase Ras plays an important role in many cellular signaling processes. Ras activity is negatively regulated by GTPase activating proteins (GAPs). It has been proposed that RasGAP may also function as an effector of Ras activity. We have identified and characterized the Drosophila homologue of the RasGAP-binding protein G3BP encoded by rasputin (rin). rin mutants are viable and display defects in photoreceptor recruitment and ommatidial polarity in the eye. Mutations in rin/G3BP genetically interact with components of the Ras signaling pathway that function at the level of Ras and above, but not with Raf/MAPK pathway components. These interactions suggest that Rin is required as an effector in Ras signaling during eye development, supporting an effector role for RasGAP. The ommatidial polarity phenotypes of rin are similar to those of RhoA and the polarity genes, e.g. fz and dsh. Although rin/G3BP interacts genetically with RhoA, affecting both photoreceptor differentiation and polarity, it does not interact with the gain-of-function genotypes of fz and dsh. These data suggest that Rin is not a general component of polarity generation, but serves a function specific to Ras and RhoA signaling pathways.

Acetylation of the c-MYC oncoprotein is required for cooperation with the HTLV-1 p30{sup II} accessory protein and the induction of oncogenic cellular transformation by p30{sup II}/c-MYC

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Romeo, Megan M.; Ko, Bookyung; Kim, Janice; Brady, Rebecca; Heatley, Hayley C.; He, Jeffrey; Harrod, Carolyn K.; Barnett, Braden [Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design, and Delivery, Southern Methodist University, Dallas, TX 75275-0376 (United States); Ratner, Lee [Departments of Medicine and Molecular Microbiology, Washington University School of Medicine, St. Louis, MO 63110 (United States); Lairmore, Michael D. [University of California-Davis, School of Veterinary Medicine, One Shields Avenue, Davis, CA 95618 (United States); Martinez, Ernest [Department of Biochemistry, University of California, Riverside, CA 92521 (United States); Lüscher, Bernhard [Institute of Biochemistry, Klinikum, RWTH Aachen University, Pauwelsstrasse 30, 52057 Aachen (Germany); Robson, Craig N. [Northern Institute for Cancer Research, Newcastle University, The Medical School, Newcastle upon Tyne, NE2 4HH (United Kingdom); Henriksson, Marie [Department of Microbiology, Cell and Tumor Biology, Karolinska Institutet, Stockholm (Sweden); Harrod, Robert, E-mail: rharrod@smu.edu [Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design, and Delivery, Southern Methodist University, Dallas, TX 75275-0376 (United States)

2015-02-15

The human T-cell leukemia retrovirus type-1 (HTLV-1) p30{sup II} protein is a multifunctional latency-maintenance factor that negatively regulates viral gene expression and deregulates host signaling pathways involved in aberrant T-cell growth and proliferation. We have previously demonstrated that p30{sup II} interacts with the c-MYC oncoprotein and enhances c-MYC-dependent transcriptional and oncogenic functions. However, the molecular and biochemical events that mediate the cooperation between p30{sup II} and c-MYC remain to be completely understood. Herein we demonstrate that p30{sup II} induces lysine-acetylation of the c-MYC oncoprotein. Acetylation-defective c-MYC Lys→Arg substitution mutants are impaired for oncogenic transformation with p30{sup II} in c-myc{sup −/−} HO15.19 fibroblasts. Using dual-chromatin-immunoprecipitations (dual-ChIPs), we further demonstrate that p30{sup II} is present in c-MYC-containing nucleoprotein complexes in HTLV-1-transformed HuT-102 T-lymphocytes. Moreover, p30{sup II} inhibits apoptosis in proliferating cells expressing c-MYC under conditions of genotoxic stress. These findings suggest that c-MYC-acetylation is required for the cooperation between p30{sup II}/c-MYC which could promote proviral replication and contribute to HTLV-1-induced carcinogenesis. - Highlights: • Acetylation of c-MYC is required for oncogenic transformation by HTLV-1 p30{sup II}/c-MYC. • Acetylation-defective c-MYC mutants are impaired for foci-formation by p30{sup II}/c-MYC. • The HTLV-1 p30{sup II} protein induces lysine-acetylation of c-MYC. • p30{sup II} is present in c-MYC nucleoprotein complexes in HTLV-1-transformed T-cells. • HTLV-1 p30{sup II} inhibits apoptosis in c-MYC-expressing proliferating cells.

Ras signaling in aging and metabolic regulation.

Science.gov (United States)

Slack, Cathy

2017-12-07

Aberrant signal transduction downstream of the Ras GTPase has a well-established role in tumorigenesis. Mutations that result in hyperactivation of Ras are responsible for a third of all human cancers. Hence, small molecule inhibitors of the Ras signal transduction cascade have been under intense focus as potential cancer treatments. In both invertebrate and mammalian models, emerging evidence has also implicated components of the Ras signaling pathway in aging and metabolic regulation. Here, I review the current evidence for Ras signaling in these newly discovered roles highlighting the interactions between the Ras pathway and other longevity assurance mechanisms. Defining the role of Ras signaling in maintaining age-related health may have important implications for the development of interventions that could not only increase lifespan but also delay the onset and/or progression of age-related functional decline.

Myc and Ras oncogenes engage different energy metabolism programs and evoke distinct patterns of oxidative and DNA replication stress

Czech Academy of Sciences Publication Activity Database

Maya-Mendoza, A.; Ostrakova, J.; Košař, Martin; Hall, A.; Duskova, P.; Mistrik, M.; Merchut-Maya, J.M.; Hodný, Zdeněk; Bartkova, J.; Christensen, C.; Bartek, Jiří

2015-01-01

Roč. 9, č. 3 (2015), s. 601-616 ISSN 1574-7891 R&D Projects: GA ČR GA13-17555S; GA MŠk(CZ) LO1304 Grant - others:Danish Council for Independent Research(DK) DFF- 1331-00262; Lundbeck Foundation(DK) R93-A8990 Institutional support: RVO:68378050 Keywords : Myc * Ras * Replication stress * DNA fork progression * Energy metabolism * DNA damage response Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.367, year: 2015

A microRNA activity map of human mesenchymal tumors: connections to oncogenic pathways; an integrative transcriptomic study

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Fountzilas Elena

2012-07-01

Full Text Available Abstract Background MicroRNAs (miRNAs are nucleic acid regulators of many human mRNAs, and are associated with many tumorigenic processes. miRNA expression levels have been used in profiling studies, but some evidence suggests that expression levels do not fully capture miRNA regulatory activity. In this study we integrate multiple gene expression datasets to determine miRNA activity patterns associated with cancer phenotypes and oncogenic pathways in mesenchymal tumors – a very heterogeneous class of malignancies. Results Using a computational method, we identified differentially activated miRNAs between 77 normal tissue specimens and 135 sarcomas and we validated many of these findings with microarray interrogation of an independent, paraffin-based cohort of 18 tumors. We also showed that miRNA activity is imperfectly correlated with miRNA expression levels. Using next-generation miRNA sequencing we identified potential base sequence alterations which may explain differential activity. We then analyzed miRNA activity changes related to the RAS-pathway and found 21 miRNAs that switch from silenced to activated status in parallel with RAS activation. Importantly, nearly half of these 21 miRNAs were predicted to regulate integral parts of the miRNA processing machinery, and our gene expression analysis revealed significant reductions of these transcripts in RAS-active tumors. These results suggest an association between RAS signaling and miRNA processing in which miRNAs may attenuate their own biogenesis. Conclusions Our study represents the first gene expression-based investigation of miRNA regulatory activity in human sarcomas, and our findings indicate that miRNA activity patterns derived from integrated transcriptomic data are reproducible and biologically informative in cancer. We identified an association between RAS signaling and miRNA processing, and demonstrated sequence alterations as plausible causes for differential miRNA activity

Oncogenes, radiation and cancer

International Nuclear Information System (INIS)

Michelin, S.C.

1998-01-01

The discovery of the oncogenic virus and the analysis of its nucleic acid, together with the development of new biochemical technology have permitted the partial knowledge of the molecular mechanisms responsible for the cellular neoplastic transformation. This work, besides describing the discovery of the first oncogenic virus and the experiments to demonstrate the existence of the oncogenes, summarizes its activation mechanisms and its intervention in cellular metabolisms. Ionizing radiation is among the external agents that induce the neoplastic process. Its participation in the genesis of this process and the contribution of oncogenes to the cellular radioresistance are among the topics, which are referred to another topic that makes reference. At the same time as the advancement of theoretical knowledge, lines of investigation for the application of the new concepts in diagnosis, prognosis and therapeutical treatment, were developed. An example of this, is the study of the participation of the oncogen c-erbB-2 in human breast cancer and its implications on the anti tumoral therapy. (author) [es

Processing and fatty acid acylation of RAS1 and RAS2 proteins in Saccharomyces cerevisiae

International Nuclear Information System (INIS)

Fujiyama, A.; Tamanoi, F.

1986-01-01

The authors demonstrate the pathway for the biosynthesis of RAS1 and RAS2 gene products of Saccharomyces cerevisiae leading to their localization in membranes. The primary translation products of these genes are detected in a soluble fraction. Shortly after synthesis, these precursor molecules are converted to forms that migrate slightly faster than the precursor forms on a NaDodSO 4 /polyacrylamide gel. These processed proteins are further modified by fatty acid acylation, which is detected by [ 3 H]palmitic acid labeling. The acylated derivatives are found exclusively in cell membranes, indicating the translocation of the RAS proteins from cytosol to membranes during maturation process. The attached fatty acids can be released by mild alkaline hydrolysis, suggesting that the linkage between the fatty acid and the protein is an ester bond. The site of the modification by fatty acid is presumably localized to the COOH-terminal portion of the RAS proteins. Fraction of the membranes by sucrose gradient demonstrates that a majority of the fatty-acylated RAS proteins are localized in plasma membrane

Ras p21 and other Gn proteins are detected in mammalian cell lines by [gamma-35S]GTP gamma S binding

International Nuclear Information System (INIS)

Comerford, J.G.; Gibson, J.R.; Dawson, A.P.; Gibson, I.

1989-01-01

The presence of guanine nucleotide binding proteins in mouse and human cell lines was investigated using [gamma- 35 S]GTP gamma S and [gamma-32P]GTP. Cell lysate polypeptides were separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis and transferred to nitrocellulose. Incubation of the nitrocellulose blots with [gamma- 35 S]GTP gamma S identified 9 distinct GTP-binding polypeptides in all lysates. One of these is the ras oncogene product, p21, as demonstrated by subsequent immunochemical staining of the nitrocellulose blots. We have shown that this procedure provides a sensitive method for detection of p21 in culture cell lines

Systems biology modeling reveals a possible mechanism of the tumor cell death upon oncogene inactivation in EGFR addicted cancers.

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Jian-Ping Zhou

Full Text Available Despite many evidences supporting the concept of "oncogene addiction" and many hypotheses rationalizing it, there is still a lack of detailed understanding to the precise molecular mechanism underlying oncogene addiction. In this account, we developed a mathematic model of epidermal growth factor receptor (EGFR associated signaling network, which involves EGFR-driving proliferation/pro-survival signaling pathways Ras/extracellular-signal-regulated kinase (ERK and phosphoinositol-3 kinase (PI3K/AKT, and pro-apoptotic signaling pathway apoptosis signal-regulating kinase 1 (ASK1/p38. In the setting of sustained EGFR activation, the simulation results show a persistent high level of proliferation/pro-survival effectors phospho-ERK and phospho-AKT, and a basal level of pro-apoptotic effector phospho-p38. The potential of p38 activation (apoptotic potential due to the elevated level of reactive oxygen species (ROS is largely suppressed by the negative crosstalk between PI3K/AKT and ASK1/p38 pathways. Upon acute EGFR inactivation, the survival signals decay rapidly, followed by a fast increase of the apoptotic signal due to the release of apoptotic potential. Overall, our systems biology modeling together with experimental validations reveals that inhibition of survival signals and concomitant release of apoptotic potential jointly contribute to the tumor cell death following the inhibition of addicted oncogene in EGFR addicted cancers.

An orthosteric inhibitor of the RAS-SOS interaction.

Science.gov (United States)

Nickerson, Seth; Joy, Stephen T; Arora, Paramjit S; Bar-Sagi, Dafna

2013-01-01

Rat sarcoma (RAS) proteins are signaling nodes that transduce extracellular cues into precise alterations in cellular physiology by engaging effector pathways. RAS signaling thus regulates diverse cell processes including proliferation, migration, differentiation, and survival. Owing to this central role in governing mitogenic signals, RAS pathway components are often dysregulated in human diseases. Targeted therapy of RAS pathways has generally not been successful, largely because of the robust biochemistry of the targets and their multifaceted network of molecular regulators. The rate-limiting step of RAS activation is Son of Sevenless (SOS)-mediated nucleotide exchange involving a single evolutionarily conserved catalytic helix from SOS. Structure function data of this mechanism provided a strong platform to design an SOS-derived, helically constrained peptide mimic as an inhibitor of the RAS-SOS interaction. In this chapter, we review RAS-SOS signaling dynamics and present evidence supporting the novel paradigm of inhibiting their interaction as a therapeutic strategy. We then describe a method of generating helically constrained peptide mimics of protein surfaces, which we have employed to inhibit the RAS-SOS active site interaction. The biochemical and functional properties of this SOS mimic support the premise that inhibition of RAS-nucleotide exchange can effectively block RAS activation and downstream signaling. © 2013 Elsevier Inc. All rights reserved.

RAS - Screens & Assays - Drug Discovery

Science.gov (United States)

The RAS Drug Discovery group aims to develop assays that will reveal aspects of RAS biology upon which cancer cells depend. Successful assay formats are made available for high-throughput screening programs to yield potentially effective drug compounds.

MECP2 Is a Frequently Amplified Oncogene with a Novel Epigenetic Mechanism That Mimics the Role of Activated RAS in Malignancy

DEFF Research Database (Denmark)

Neupane, Manish; Clark, Allison P.; Landini, Serena

2016-01-01

An unbiased genome-scale screen for unmutated genes that drive cancer growth when overexpressed identified methyl cytosine-guanine dinucleotide (CpG) binding protein 2 (MECP2) as a novel oncogene. MECP2 resides in a region of the X-chromosome that is significantly amplified across 18% of cancers,...

A genetic screen for anchorage-independent proliferation in mammalian cells identifies a membrane-bound neuregulin.

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Davide Danovi

2010-07-01

Full Text Available Anchorage-independent proliferation is a hallmark of oncogenic transformation and is thought to be conducive to proliferation of cancer cells away from their site of origin. We have previously reported that primary Schwann cells expressing the SV40 Large T antigen (LT are not fully transformed in that they maintain a strict requirement for attachment, requiring a further genetic change, such as oncogenic Ras, to gain anchorage-independence. Using the LT-expressing cells, we performed a genetic screen for anchorage-independent proliferation and identified Sensory and Motor Neuron Derived Factor (SMDF, a transmembrane class III isoform of Neuregulin 1. In contrast to oncogenic Ras, SMDF induced enhanced proliferation in normal primary Schwann cells but did not trigger cellular senescence. In cooperation with LT, SMDF drove anchorage-independent proliferation, loss of contact inhibition and tumourigenicity. This transforming ability was shared with membrane-bound class III but not secreted class I isoforms of Neuregulin, indicating a distinct mechanism of action. Importantly, we show that despite being membrane-bound signalling molecules, class III neuregulins transform via a cell intrinsic mechanism, as a result of constitutive, elevated levels of ErbB signalling at high cell density and in anchorage-free conditions. This novel transforming mechanism may provide new targets for cancer therapy.

Tyrosine 842 in the activation loop is required for full transformation by the oncogenic mutant FLT3-ITD.

Science.gov (United States)

Kazi, Julhash U; Chougule, Rohit A; Li, Tianfeng; Su, Xianwei; Moharram, Sausan A; Rupar, Kaja; Marhäll, Alissa; Gazi, Mohiuddin; Sun, Jianmin; Zhao, Hui; Rönnstrand, Lars

2017-07-01

The type III receptor tyrosine kinase FLT3 is frequently mutated in acute myeloid leukemia. Oncogenic FLT3 mutants display constitutive activity leading to aberrant cell proliferation and survival. Phosphorylation on several critical tyrosine residues is known to be essential for FLT3 signaling. Among these tyrosine residues, Y842 is located in the so-called activation loop. The position of this tyrosine residue is well conserved in all receptor tyrosine kinases. It has been reported that phosphorylation of the activation loop tyrosine is critical for catalytic activity for some but not all receptor tyrosine kinases. The role of Y842 residue in FLT3 signaling has not yet been studied. In this report, we show that Y842 is not important for FLT3 activation or ubiquitination but plays a critical role in regulating signaling downstream of the receptor as well as controlling receptor stability. We found that mutation of Y842 in the FLT3-ITD oncogenic mutant background reduced cell viability and increased apoptosis. Furthermore, the introduction of the Y842 mutation in the FLT3-ITD background led to a dramatic reduction in in vitro colony forming capacity. Additionally, mice injected with cells expressing FLT3-ITD/Y842F displayed a significant delay in tumor formation, compared to FLT3-ITD expressing cells. Microarray analysis comparing gene expression regulated by FLT3-ITD versus FLT3-ITD/Y842F demonstrated that mutation of Y842 causes suppression of anti-apoptotic genes. Furthermore, we showed that cells expressing FLT3-ITD/Y842F display impaired activity of the RAS/ERK pathway due to reduced interaction between FLT3 and SHP2 leading to reduced SHP2 activation. Thus, we suggest that Y842 is critical for FLT3-mediated RAS/ERK signaling and cellular transformation.

Analog reactor simulator RAS; Reaktorski analogni simulator RAS

Energy Technology Data Exchange (ETDEWEB)

Radanovic, Lj; Bingulac, S; Popovic, D [The Institute of Nuclear Sciences Boris Kidric, Vinca, Beograd (Yugoslavia)

1961-07-01

Analog computer RAS was designed as a nuclear reactor simulator, but it can be simultaneously used for solving a number of other problems. This paper contains a brief description of the simulator parts and their principal characteristics.

Using the capital markets in Ras Gas

International Nuclear Information System (INIS)

Voge, B.; Penzer, M.

1997-01-01

In December 1996, Ras Laffan Liquefied Natural Gas Company Ltd (Ras Gas) closed a multi-source financing that included an offering of US$1.2bn of bonds. The sponsors of the Ras Gas project overcame a number of obstacles on the road to closing the capital markets offering. This article provides a general overview of capital markets offerings in international project financings and discusses how Ras Gas was able to successfully integrate a capital markets offering into a financing plan which included a commercial bank facility and several export-credit agency facilities. (Author)

Inhibition of RAS in diabetic nephropathy

Directory of Open Access Journals (Sweden)

Yacoub R

2015-04-01

Full Text Available Rabi Yacoub, Kirk N Campbell Department of Medicine, Division of Nephrology, Icahn School of Medicine at Mount Sinai, New York, NY, USA Abstract: Diabetic kidney disease (DKD is a progressive proteinuric renal disorder in patients with type 1 or type 2 diabetes mellitus. It is a common cause of end-stage kidney disease worldwide, particularly in developed countries. Therapeutic targeting of the renin–angiotensin system (RAS is the most validated clinical strategy for slowing disease progression. DKD is paradoxically a low systematic renin state with an increased intrarenal RAS activity implicated in its pathogenesis. Angiotensin II (AngII, the main peptide of RAS, is not only a vasoactive peptide but functions as a growth factor, activating interstitial fibroblasts and mesangial and tubular cells, while promoting the synthesis of extracellular matrix proteins. AngII also promotes podocyte injury through increased calcium influx and the generation of reactive oxygen species. Blockade of the RAS using either angiotensin converting enzyme inhibitors, or angiotensin receptor blockers can attenuate progressive glomerulosclerosis in animal models, and slows disease progression in humans with DKD. In this review, we summarize the role of intrarenal RAS activation in the pathogenesis and progression of DKD and the rationale for RAS inhibition in this population. Keywords: renin–angiotensin system, diabetic kidney disease, angiotensin II, angiotensin-converting enzyme inhibitors, angiotensin receptor blockers

BRAF and RAS oncogenes regulate Rho GTPase pathways to mediate migration and invasion properties in human colon cancer cells: a comparative study

Czech Academy of Sciences Publication Activity Database

Makrodouli, E.; Oikonomou, E.; Koc, Michal; Anděra, Ladislav; Sasazuki, T.; Shirasawa, S.; Pintzas, A.

2011-01-01

Roč. 10, - (2011), e118 ISSN 1476-4598 Grant - others:GSRT(GR) 03ED562; EK(XE) LSHC-CT-2006-037278 Institutional research plan: CEZ:AV0Z50520514 Keywords : colorectal cancer * invasiveness * oncogenes Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.993, year: 2011

Activated H-Ras regulates hematopoietic cell survival by modulating Survivin

International Nuclear Information System (INIS)

Fukuda, Seiji; Pelus, Louis M.

2004-01-01

Survivin expression and Ras activation are regulated by hematopoietic growth factors. We investigated whether activated Ras could circumvent growth factor-regulated Survivin expression and if a Ras/Survivin axis mediates growth factor independent survival and proliferation in hematopoietic cells. Survivin expression is up-regulated by IL-3 in Ba/F3 and CD34 + cells and inhibited by the Ras inhibitor, farnesylthiosalicylic acid. Over-expression of constitutively activated H-Ras (CA-Ras) in Ba/F3 cells blocked down-modulation of Survivin expression, G 0 /G 1 arrest, and apoptosis induced by IL-3 withdrawal, while dominant-negative (DN) H-Ras down-regulated Survivin. Survivin disruption by DN T34A Survivin blocked CA-Ras-induced IL-3-independent cell survival and proliferation; however, it did not affect CA-Ras-mediated enhancement of S-phase, indicating that the anti-apoptotic activity of CA-Ras is Survivin dependent while its S-phase enhancing effect is not. These results indicate that CA-Ras modulates Survivin expression independent of hematopoietic growth factors and that a CA-Ras/Survivin axis regulates survival and proliferation of transformed hematopoietic cells

Renal Artery Stenosis (RAS) Case study

International Nuclear Information System (INIS)

Zaater, M.K.

2012-01-01

Renal Artery Stenosis (RAS), is one of the causes of secondary hypertension; there are many causes of renal artery stenosis, as atherosclerosis of the renal artery which account for 90% of cases of RAS; fibromuscular dysplasia accounts for 10% of RAS. Various causes of thrombophilia either due congenital causes or acquired causes and can lead to RAS. Our patient was presented by acute attack of epistaxis and hypertension. Angiography of the Renal Arteries,are showed no sign of renal artery stenosis. However, the right kidney showed upper pole infarction, and the left kidney showed evidence of functional lower pole renal artery stenosis, although there is no anatomical stenosis detected in angiography. Work up for the cause of thrombophilia did not help in the diagnosis, which may be due to an undiscovered cause of thrombophilia

Coupling between p210bcr-abl and Shc and Grb2 adaptor proteins in hematopoietic cells permits growth factor receptor-independent link to ras activation pathway.

Science.gov (United States)

Tauchi, T; Boswell, H S; Leibowitz, D; Broxmeyer, H E

1994-01-01

Enforced expression of p210bcr-abl transforms interleukin 3 (IL-3)-dependent hematopoietic cell lines to growth factor-independent proliferation. It has been demonstrated that nonreceptor tyrosine kinase oncogenes may couple to the p21ras pathway to exert their transforming effect. In particular, p210bcr-abl was recently found to effect p21ras activation in hematopoietic cells. In this context, experiments were performed to evaluate a protein signaling pathway by which p210bcr-abl might regulate p21ras. It was asked whether Shc p46/p52, a protein containing a src-homology region 2 (SH2) domain, and known to function upstream from p21ras, might form specific complexes with p210bcr-abl and thus, possibly alter p21ras activity by coupling to the guanine nucleotide exchange factor (Sos/CDC25) through the Grb2 protein-Sos complex. This latter complex has been previously demonstrated to occur ubiquitously. We found that p210bcr-abl formed a specific complex with Shc and with Grb2 in three different murine cell lines transfected with a p210bcr-abl expression vector. There appeared to be a higher order complex containing Shc, Grb2, and bcr-abl proteins. In contrast to p210bcr-abl transformed cells, in which there was constitutive tight association between Grb2 and Shc, binding between Grb2 and Shc was Steel factor (SLF)-dependent in a SLF-responsive, nontransformed parental cell line. The SLF-dependent association between Grb2 and Shc in nontransformed cells involved formation of a complex of Grb2 with c-kit receptor after SLF treatment. Thus, p210bcr-abl appears to function in a hematopoietic p21ras activation pathway to allow growth factor-independent coupling between Grb2, which exists in a complex with the guanine nucleotide exchange factor (Sos), and p21ras. Shc may not be required for Grb2-c-kit interaction, because it fails to bind strongly to c-kit.

RasGRP1, but not RasGRP3, is required for efficient thymic β-selection and ERK activation downstream of CXCR4.

Directory of Open Access Journals (Sweden)

Dominic P Golec

Full Text Available T cell development is a highly dynamic process that is driven by interactions between developing thymocytes and the thymic microenvironment. Upon entering the thymus, the earliest thymic progenitors, called CD4(-CD8(- 'double negative' (DN thymocytes, pass through a checkpoint termed "β-selection" before maturing into CD4(+CD8(+ 'double positive' (DP thymocytes. β-selection is an important developmental checkpoint during thymopoiesis where developing DN thymocytes that successfully express the pre-T cell receptor (TCR undergo extensive proliferation and differentiation towards the DP stage. Signals transduced through the pre-TCR, chemokine receptor CXCR4 and Notch are thought to drive β-selection. Additionally, it has long been known that ERK is activated during β-selection; however the pathways regulating ERK activation remain unknown. Here, we performed a detailed analysis of the β-selection events in mice lacking RasGRP1, RasGRP3 and RasGRP1 and 3. We report that RasGRP1 KO and RasGRP1/3 DKO deficient thymi show a partial developmental block at the early DN3 stage of development. Furthermore, DN3 thymocytes from RasGRP1 and RasGRP1/3 double knock-out thymi show significantly reduced proliferation, despite expression of the TCRβ chain. As a result of impaired β-selection, the pool of TCRβ(+ DN4 is significantly diminished, resulting in inefficient DN to DP development. Also, we report that RasGRP1 is required for ERK activation downstream of CXCR4 signaling, which we hypothesize represents a potential mechanism of RasGRP1 regulation of β-selection. Our results demonstrate that RasGRP1 is an important regulator of proliferation and differentiation at the β-selection checkpoint and functions downstream of CXCR4 to activate the Ras/MAPK pathway.

Identifying Breast Cancer Oncogenes

Science.gov (United States)

2011-10-01

cells we observed that it promoted transformation of HMLE cells, suggesting a tumor suppressive role of Merlin in breast cancer (Figure 4B). A...08-1-0767 TITLE: Identifying Breast Cancer Oncogenes PRINCIPAL INVESTIGATOR: Yashaswi Shrestha...Standard Form 298 (Rev. 8-98) Prescribed by ANSI Std. Z39.18 W81XWH-08-1-0767 Identifying Breast Cancer Oncogenes Yashaswi Shrestha Dana-Farber

Oncogenic cancer/testis antigens

DEFF Research Database (Denmark)

Gjerstorff, Morten F; Andersen, Mads H; Ditzel, Henrik J

2015-01-01

Recent developments have set the stage for immunotherapy as a supplement to conventional cancer treatment. Consequently, a significant effort is required to further improve efficacy and specificity, particularly the identification of optimal therapeutic targets for clinical testing. Cancer....../testis antigens are immunogenic, highly cancer-specific, and frequently expressed in various types of cancer, which make them promising candidate targets for cancer immunotherapy, including cancer vaccination and adoptive T-cell transfer with chimeric T-cell receptors. Our current understanding of tumor...... immunology and immune escape suggests that targeting oncogenic antigens may be beneficial, meaning that identification of cancer/testis antigens with oncogenic properties is of high priority. Recent work from our lab and others provide evidence that many cancer/testis antigens, in fact, have oncogenic...

Ras and Rheb Signaling in Survival and Cell Death

International Nuclear Information System (INIS)

Ehrkamp, Anja; Herrmann, Christian; Stoll, Raphael; Heumann, Rolf

2013-01-01

One of the most obvious hallmarks of cancer is uncontrolled proliferation of cells partly due to independence of growth factor supply. A major component of mitogenic signaling is Ras, a small GTPase. It was the first identified human protooncogene and is known since more than three decades to promote cellular proliferation and growth. Ras was shown to support growth factor-independent survival during development and to protect from chemical or mechanical lesion-induced neuronal degeneration in postmitotic neurons. In contrast, for specific patho-physiological cases and cellular systems it has been shown that Ras may also promote cell death. Proteins from the Ras association family (Rassf, especially Rassf1 and Rassf5) are tumor suppressors that are activated by Ras-GTP, triggering apoptosis via e.g., activation of mammalian sterile 20-like (MST1) kinase. In contrast to Ras, their expression is suppressed in many types of tumours, which makes Rassf proteins an exciting model for understanding the divergent effects of Ras activity. It seems likely that the outcome of Ras signaling depends on the balance between the activation of its various downstream effectors, thus determining cellular fate towards either proliferation or apoptosis. Ras homologue enriched in brain (Rheb) is a protein from the Ras superfamily that is also known to promote proliferation, growth, and regeneration through the mammalian target of rapamycin (mTor) pathway. However, recent evidences indicate that the Rheb-mTor pathway may switch its function from a pro-growth into a cell death pathway, depending on the cellular situation. In contrast to Ras signaling, for Rheb, the cellular context is likely to modulate the whole Rheb-mTor pathway towards cellular death or survival, respectively

Ras and Rheb Signaling in Survival and Cell Death

Energy Technology Data Exchange (ETDEWEB)

Ehrkamp, Anja [Molecular Neurobiochemistry, Ruhr University of Bochum, 44780 Bochum (Germany); Herrmann, Christian [Department of Physical Chemistry1, Protein Interaction, Ruhr University of Bochum, 44780 Bochum (Germany); Stoll, Raphael [Biomolecular NMR, Ruhr University of Bochum, 44780 Bochum (Germany); Heumann, Rolf, E-mail: rolf.heumann@rub.de [Molecular Neurobiochemistry, Ruhr University of Bochum, 44780 Bochum (Germany)

2013-05-28

One of the most obvious hallmarks of cancer is uncontrolled proliferation of cells partly due to independence of growth factor supply. A major component of mitogenic signaling is Ras, a small GTPase. It was the first identified human protooncogene and is known since more than three decades to promote cellular proliferation and growth. Ras was shown to support growth factor-independent survival during development and to protect from chemical or mechanical lesion-induced neuronal degeneration in postmitotic neurons. In contrast, for specific patho-physiological cases and cellular systems it has been shown that Ras may also promote cell death. Proteins from the Ras association family (Rassf, especially Rassf1 and Rassf5) are tumor suppressors that are activated by Ras-GTP, triggering apoptosis via e.g., activation of mammalian sterile 20-like (MST1) kinase. In contrast to Ras, their expression is suppressed in many types of tumours, which makes Rassf proteins an exciting model for understanding the divergent effects of Ras activity. It seems likely that the outcome of Ras signaling depends on the balance between the activation of its various downstream effectors, thus determining cellular fate towards either proliferation or apoptosis. Ras homologue enriched in brain (Rheb) is a protein from the Ras superfamily that is also known to promote proliferation, growth, and regeneration through the mammalian target of rapamycin (mTor) pathway. However, recent evidences indicate that the Rheb-mTor pathway may switch its function from a pro-growth into a cell death pathway, depending on the cellular situation. In contrast to Ras signaling, for Rheb, the cellular context is likely to modulate the whole Rheb-mTor pathway towards cellular death or survival, respectively.

Electrostatic Interactions Positively Regulate K-Ras Nanocluster Formation and Function▿

Science.gov (United States)

Plowman, Sarah J.; Ariotti, Nicholas; Goodall, Andrew; Parton, Robert G.; Hancock, John F.

2008-01-01

The organization of Ras proteins into plasma membrane nanoclusters is essential for high-fidelity signal transmission, but whether the nanoscale enviroments of different Ras nanoclusters regulate effector interactions is unknown. We show using high-resolution spatial mapping that Raf-1 is recruited to and retained in K-Ras-GTP nanoclusters. In contrast, Raf-1 recruited to the plasma membrane by H-Ras is not retained in H-Ras-GTP nanoclusters. Similarly, upon epidermal growth factor receptor activation, Raf-1 is preferentially recruited to K-Ras-GTP and not H-Ras-GTP nanoclusters. The formation of K-Ras-GTP nanoclusters is inhibited by phosphorylation of S181 in the C-terminal polybasic domain or enhanced by blocking S181 phosphorylation, with a concomitant reduction or increase in Raf-1 plasma membrane recruitment, respectively. Phosphorylation of S181 does not, however, regulate in vivo interactions with the nanocluster scaffold galectin-3 (Gal3), indicating separate roles for the polybasic domain and Gal3 in driving K-Ras nanocluster formation. Together, these data illustrate that Ras nanocluster composition regulates effector recruitment and highlight the importance of lipid/protein nanoscale environments to the activation of signaling cascades. PMID:18458061

An oncogenic MYB feedback loop drives alternate cell fates in adenoid cystic carcinoma

Science.gov (United States)

Drier, Yotam; Cotton, Matthew J.; Williamson, Kaylyn E.; Gillespie, Shawn M.; Ryan, Russell J.H.; Kluk, Michael J.; Carey, Christopher D.; Rodig, Scott J.; Sholl, Lynette M; Afrogheh, Amir H.; Faquin, William C.; Queimado, Lurdes; Qi, Jun; Wick, Michael J.; El-Naggar, Adel K.; Bradner, James E.; Moskaluk, Christopher A.; Aster, Jon C.; Knoechel, Birgit; Bernstein, Bradley E.

2016-01-01

Translocation events are frequent in cancer and may create chimeric fusions or ‘regulatory rearrangements’ that drive oncogene overexpression. Here we identify super-enhancer translocations that drive overexpression of the oncogenic transcription factor MYB as a recurrent theme in adenoid cystic carcinoma (ACC). Whole-genome sequencing data and chromatin maps reveal distinct chromosomal rearrangements that juxtapose super-enhancers to the MYB locus. Chromosome conformation capture confirms that the translocated enhancers interact with the MYB promoter. Remarkably, MYB protein binds to the translocated enhancers, creating a positive feedback loop that sustains its expression. MYB also binds enhancers that drive different regulatory programs in alternate cell lineages in ACC, cooperating with TP63 in myoepithelial cells and a Notch program in luminal epithelial cells. Bromodomain inhibitors slow tumor growth in ACC primagraft models in vivo. Thus, our study identifies super-enhancer translocations that drive MYB expression and provides insight into downstream MYB functions in the alternate ACC lineages. PMID:26829750

Flavopiridol Synergizes with Sorafenib to Induce Cytotoxicity and Potentiate Antitumorigenic Activity in EGFR/HER-2 and Mutant RAS/RAF Breast Cancer Model Systems

Directory of Open Access Journals (Sweden)

Teddy S Nagaria

2013-08-01

Full Text Available Oncogenic receptor tyrosine kinase (RTK signaling through the Ras-Raf-Mek-Erk (Ras-MAPK pathway is implicated in a wide array of carcinomas, including those of the breast. The cyclin-dependent kinases (CDKs are implicated in regulating proliferative and survival signaling downstream of this pathway. Here, we show that CDK inhibitors exhibit an order of magnitude greater cytotoxic potency than a suite of inhibitors targeting RTK and Ras-MAPK signaling in cell lines representative of clinically recognized breast cancer (BC subtypes. Drug combination studies show that the pan-CDK inhibitor, flavopiridol (FPD, synergistically potentiated cytotoxicity induced by the Raf inhibitor, sorafenib (SFN. This synergy was most pronounced at sub-EC50 SFN concentrations in MDA-MB-231 (KRAS-G13D and BRAF-G464V mutations, MDA-MB-468 [epidermal growth factor receptor (EGFR overexpression], and SKBR3 [ErbB2/EGFR2 (HER-2 overexpression] cells but not in hormone-dependent MCF-7 and T47D cells. Potentiation of SFN cytotoxicity by FPD correlated with enhanced apoptosis, suppression of retinoblastoma (Rb signaling, and reduced Mcl-1 expression. SFN and FPD were also tested in an MDA-MB-231 mammary fat pad engraftment model of tumorigenesis. Mice treated with both drugs exhibited reduced primary tumor growth rates and metastatic tumor load in the lungs compared to treatment with either drug alone, and this correlated with greater reductions in Rb signaling and Mcl-1 expression in resected tumors. These findings support the development of CDK and Raf co-targeting strategies in EGFR/HER-2-overexpressing or RAS/RAF mutant BCs.

Interaction of CtBP with adenovirus E1A suppresses immortalization of primary epithelial cells and enhances virus replication during productive infection.

Science.gov (United States)

Subramanian, T; Zhao, Ling-Jun; Chinnadurai, G

2013-09-01

Adenovirus E1A induces cell proliferation, oncogenic transformation and promotes viral replication through interaction with p300/CBP, TRRAP/p400 multi-protein complex and the retinoblastoma (pRb) family proteins through distinct domains in the E1A N-terminal region. The C-terminal region of E1A suppresses E1A/Ras co-transformation and interacts with FOXK1/K2, DYRK1A/1B/HAN11 and CtBP1/2 (CtBP) protein complexes. To specifically dissect the role of CtBP interaction with E1A, we engineered a mutation (DL→AS) within the CtBP-binding motif, PLDLS, and investigated the effect of the mutation on immortalization and Ras cooperative transformation of primary cells and viral replication. Our results suggest that CtBP-E1A interaction suppresses immortalization and Ras co-operative transformation of primary rodent epithelial cells without significantly influencing the tumorigenic activities of transformed cells in immunodeficient and immunocompetent animals. During productive infection, CtBP-E1A interaction enhances viral replication in human cells. Between the two CtBP family proteins, CtBP2 appears to restrict viral replication more than CtBP1 in human cells. Copyright © 2013 Elsevier Inc. All rights reserved.

Ras and relatives--job sharing and networking keep an old family together.

Science.gov (United States)

Ehrhardt, Annette; Ehrhardt, Götz R A; Guo, Xuecui; Schrader, John W

2002-10-01

Many members of the Ras superfamily of GTPases have been implicated in the regulation of hematopoietic cells, with roles in growth, survival, differentiation, cytokine production, chemotaxis, vesicle-trafficking, and phagocytosis. The well-known p21 Ras proteins H-Ras, N-Ras, K-Ras 4A, and K-Ras 4B are also frequently mutated in human cancer and leukemia. Besides the four p21 Ras proteins, the Ras subfamily of the Ras superfamily includes R-Ras, TC21 (R-Ras2), M-Ras (R-Ras3), Rap1A, Rap1B, Rap2A, Rap2B, RalA, and RalB. They exhibit remarkable overall amino acid identities, especially in the regions interacting with the guanine nucleotide exchange factors that catalyze their activation. In addition, there is considerable sharing of various downstream effectors through which they transmit signals and of GTPase activating proteins that downregulate their activity, resulting in overlap in their regulation and effector function. Relatively little is known about the physiological functions of individual Ras family members, although the presence of well-conserved orthologs in Caenorhabditis elegans suggests that their individual roles are both specific and vital. The structural and functional similarities have meant that commonly used research tools fail to discriminate between the different family members, and functions previously attributed to one family member may be shared with other members of the Ras family. Here we discuss similarities and differences in activation, effector usage, and functions of different members of the Ras subfamily. We also review the possibility that the differential localization of Ras proteins in different parts of the cell membrane may govern their responses to activation of cell surface receptors.

An Oncogenic Role for Alternative NF-κB Signaling in DLBCL Revealed upon Deregulated BCL6 Expression

Directory of Open Access Journals (Sweden)

Baochun Zhang

2015-05-01

Full Text Available Diffuse large B cell lymphoma (DLBCL is a complex disease comprising diverse subtypes and genetic profiles. Possibly because of the prevalence of genetic alterations activating canonical NF-κB activity, a role for oncogenic lesions that activate the alternative NF-κB pathway in DLBCL has remained elusive. Here, we show that deletion/mutation of TRAF3, a negative regulator of the alternative NF-κB pathway, occurs in ∼15% of DLBCLs and that it often coexists with BCL6 translocation, which prevents terminal B cell differentiation. Accordingly, in a mouse model constitutive activation of the alternative NF-κB pathway cooperates with BCL6 deregulation in DLBCL development. This work demonstrates a key oncogenic role for the alternative NF-κB pathway in DLBCL development.

Two-dimensional electrophoretic analysis of transformation-sensitive polypeptides during chemically, spontaneously, and oncogene-induced transformation of rat liver epithelial cells

DEFF Research Database (Denmark)

Wirth, P J; Luo, L D; Fujimoto, Y

1992-01-01

; AFB), spontaneously, and oncogene (v-Ha-ras, v-raf, and v-myc/v-raf)-induced transformation of RLE cells. Two-dimensional mapping of [35S]methionine-labeled whole cell lysate, cell-free in vitro translation products and [32P]orthophosphate-labeled polypeptides revealed subsets of polypeptides specific...... for each transformation modality. A search of the RLE protein database indicated the specific subcellular location for the majority of these transformation-sensitive proteins. Significant alterations in the expression of the extracellular matrix protein, fibronectin, as well as tropomyosin......- and intermediate filament-related polypeptides (vimentin, beta-tubulin, the cytokeratins, and actin) were observed among the various transformant cell lines. Immunoprecipitation and Western immunoblot analysis of tropomyosin expression in four individual AFB-, as well as four spontaneously induced, and each...

Mutational analysis of a ras catalytic domain

DEFF Research Database (Denmark)

Willumsen, B M; Papageorge, A G; Kung, H F

1986-01-01

localization. We speculate that this latter region interacts with the putative cellular target of ras. The results suggest that transforming ras proteins require membrane localization, guanosine nucleotide binding, and an additional undefined function that may represent interaction with their target....

Interplay among Drosophila transcription factors Ets21c, Fos and Ftz-F1 drives JNK-mediated tumor malignancy

Directory of Open Access Journals (Sweden)

Eva Külshammer

2015-10-01

Full Text Available Cancer initiation and maintenance of the transformed cell state depend on altered cellular signaling and aberrant activities of transcription factors (TFs that drive pathological gene expression in response to cooperating genetic lesions. Deciphering the roles of interacting TFs is therefore central to understanding carcinogenesis and for designing cancer therapies. Here, we use an unbiased genomic approach to define a TF network that triggers an abnormal gene expression program promoting malignancy of clonal tumors, generated in Drosophila imaginal disc epithelium by gain of oncogenic Ras (RasV12 and loss of the tumor suppressor Scribble (scrib1. We show that malignant transformation of the rasV12scrib1 tumors requires TFs of distinct families, namely the bZIP protein Fos, the ETS-domain factor Ets21c and the nuclear receptor Ftz-F1, all acting downstream of Jun-N-terminal kinase (JNK. Depleting any of the three TFs improves viability of tumor-bearing larvae, and this positive effect can be enhanced further by their combined removal. Although both Fos and Ftz-F1 synergistically contribute to rasV12scrib1 tumor invasiveness, only Fos is required for JNK-induced differentiation defects and Matrix metalloprotease (MMP1 upregulation. In contrast, the Fos-dimerizing partner Jun is dispensable for JNK to exert its effects in rasV12scrib1 tumors. Interestingly, Ets21c and Ftz-F1 are transcriptionally induced in these tumors in a JNK- and Fos-dependent manner, thereby demonstrating a hierarchy within the tripartite TF network, with Fos acting as the most upstream JNK effector. Of the three TFs, only Ets21c can efficiently substitute for loss of polarity and cooperate with RasV12 in inducing malignant clones that, like rasV12scrib1 tumors, invade other tissues and overexpress MMP1 and the Drosophila insulin-like peptide 8 (Dilp8. While rasV12ets21c tumors require JNK for invasiveness, the JNK activity is dispensable for their growth. In conclusion, our

The oncogenic action of ionizing radiation on rat skin: Progress report, February 1, 1987-January 31, 1988

International Nuclear Information System (INIS)

Burns, F.J.

1987-01-01

The work outlined in this report includes: epidermal DNA strand breaks and radiation penetration; activation of oncogenes in radiation induced rat skin tumors; and rat skin carcinogenesis by neon ions. The effect of radiation penetration on DNA single strand breaks has been studied extensively in rat and mouse epidermis. The results show clearly that the number of strand breaks per unit dose in the epidermal DNA is reduced by 50% to 65% when the radiation penetration is reduced from 1.0 mm to 0.2 mm. This penetration effect on DNA strand breaks was not seen in mouse epidermal cell lines growing in plastic dishes. The results imply that DNA strand breakage in superficial cells is partially dependent on the radiation dose to underlying tissue. The phenomenon is not mediated by systemic interactions as it was observed in irradiated explanted skin. The oncogene activation pattern in the radiation-induced skin tumors was found to be tumor dependent. Either K-ras activation or c-myc amplification or both was observed in each tumor analyzed so far. Even benign fibromas exhibited c-myc amplification. The carcinogenicity of high penetration electrons (2.0 MeV) was determined in preparation for similar studies with a neon ion beam at the Berkeley Bevelac. The principal finding so far is a large excess of connective tissue tumors, fibromas (benign) and sarcomas (malignant). 59 refs., 1 tab

Net (ERP/SAP2) one of the Ras-inducible TCFs, has a novel inhibitory domain with resemblance to the helix-loop-helix motif.

Science.gov (United States)

Maira, S M; Wurtz, J M; Wasylyk, B

1996-11-01

The three ternary complex factors (TCFs), Net (ERP/ SAP-2), ELK-1 and SAP-1, are highly related ets oncogene family members that participate in the response of the cell to Ras and growth signals. Understanding the different roles of these factors will provide insights into how the signals result in coordinate regulation of the cell. We show that Net inhibits transcription under basal conditions, in which SAP-1a is inactive and ELK-1 stimulates. Repression is mediated by the NID, the Net Inhibitory Domain of about 50 amino acids, which autoregulates the Net protein and also inhibits when it is isolated in a heterologous fusion protein. Net is particularly sensitive to Ras activation. Ras activates Net through the C-domain, which is conserved between the three TCFs, and the NID is an efficient inhibitor of Ras activation. The NID, as well as more C-terminal sequences, inhibit DNA binding. Net is more refractory to DNA binding than the other TCFs, possibly due to the presence of multiple inhibitory elements. The NID may adopt a helix-loop-helix (HLH) structure, as evidenced by homology to other HLH motifs, structure predictions, model building and mutagenesis of critical residues. The sequence resemblance with myogenic factors suggested that Net may form complexes with the same partners. Indeed, we found that Net can interact in vivo with the basic HLH factor, E47. We propose that Net is regulated at the level of its latent DNA-binding activity by protein interactions and/or phosphorylation. Net may form complexes with HLH proteins as well as SRF on specific promotor sequences. The identification of the novel inhibitory domain provides a new inroad into exploring the different roles of the ternary complex factors in growth control and transformation.

Regional cooperation in RWM in the AO countries

International Nuclear Information System (INIS)

Ya-anant, N.; Thiangtrongjit, S.

2011-01-01

Regional cooperation in radioactive waste management (RWM) in the Asian and Oceanic countries has been successfully developing for more than 20 y. At present, the Asian and Oceanic countries recognise that there are several projects under the regional cooperation, such as the Forum for Nuclear Cooperation in Asia (FNCA), the Asian Nuclear Safety Network (ANSN) and the International Atomic Energy Agency (IAEA) regional projects. The FNCA on Radiation Safety and RWM is an effective mechanism for enhancing socio-economic development through active regional partnership in the peaceful and safe utilisation of nuclear technology. The ANSN Topical Group on RWM is established for exchanging, pooling, and sharing knowledge in the field of nuclear safety. For IAEA regional cooperation on RWM, such as IAEA-regional project RAS/3/009 Strengthening Infrastructure of RWM can provide and support Asian and Oceanic countries for national work plan, fellowships and scientific visits as well as expert services. (authors)

Opposite effects of Ha-Ras and Ki-Ras on radiation-induced apoptosis via differential activation of PI3K/Akt and Rac/p38 mitogen-activated protein kinase signaling

International Nuclear Information System (INIS)

Choi, J.-A.; Kang, C.-M.; Lee, Y.-S.; Lee, S.-J.; Bae, S.-W.; Cho, C.-K.

2003-01-01

It has been well known that Ras signaling is involved in various cellular processes, including proliferation, differentiation, and apoptosis. However, distinct cellular functions of Ras isozymes are not fully understood. Here we show the opposing roles of Ha-Ras and Ki-Ras genes in the modulation of cell sensitivity to ionizing radiation. Overexpression of active isoform of Ha-Ras (12V-Ha- Ras) in Rat2 cells increases resistance to the ionizing radiation. Constitutive activation of phosphoinositide-3-kinase (PI3K) and Akt is detected specifically in 12V-Ha-Ras-overexpressing cells. The specific PI3K inhibitor LY294002 inhibits PI3K/Akt signaling and potentiates the radiation-induced apoptosis, suggesting that activation of PI3K/Akt signaling pathway is involved in the increased radio-resistance in cells overexpressing 12V-Ha-Ras. Overexpression of activated Ki-Ras (12V-Ki-Ras), on the other hand, markedly increases radiation sensitivity. The p38 mitogen-activated protein (MAP) kinase activity is selectively enhanced by ionizing radiation in cells overexpressing 12V-Ki-Ras. The specific p38 MAP kinase inhibitor, PD169316, or dominant-negative p38 MAP kinase decreases radiation-induced cell death. We further show that the mechanism that underlies potentiation of cell death in cells overexpressing 12V-Ki-Ras involves Bax translocation to the mitochondrial membrane. Elevated Bax translocation following ionizing irradiation in 12V-Ki-Ras-overexpressing cells is completely inhibited by PD169316 or dominant-negative p38 MAP kinase. In addition, introduction of cells with RacN17, a dominant negative mutant of Rac, resulted in a marked inhibition of radiation-induced Bax translocation and apoptotic cell death as well as p38 MAP kinase activation. Taken together, these findings explain the opposite effects of Ha-Ras and Ki-Ras on modulation of radio-sensitivity, and suggest that differential activation of PI3K/Akt and Rac/p38 MAP kinase signaling by Ha-Ras and Ki-Ras may

RasGRP3 regulates the migration of glioma cells via interaction with Arp3

Science.gov (United States)

Lee, Hae Kyung; Finniss, Susan; Cazacu, Simona; Xiang, Cunli; Poisson, Laila M.; Blumberg, Peter M.; Brodie, Chaya

2015-01-01

Glioblastoma (GBM), the most aggressive primary brain tumors, are highly infiltrative. Although GBM express high Ras activity and Ras proteins have been implicated in gliomagenesis, Ras-activating mutations are not frequent in these tumors. RasGRP3, an important signaling protein responsive to diacylglycerol (DAG), increases Ras activation. Here, we examined the expression and functions of RasGRP3 in GBM and glioma cells. RasGRP3 expression was upregulated in GBM specimens and glioma stem cells compared with normal brains and neural stem cells, respectively. RasGRP3 activated Ras and Rap1 in glioma cells and increased cell migration and invasion partially via Ras activation. Using pull-down assay and mass spectroscopy we identified the actin-related protein, Arp3, as a novel interacting protein of RasGRP3. The interaction of RasGRP3 and Arp3 was validated by immunofluorescence staining and co-immunoprecipitation, and PMA, which activates RasGRP3 and induces its translocation to the peri-nuclear region, increased the association of Arp3 and RasGRP3. Arp3 was upregulated in GBM, regulated cell spreading and migration and its silencing partially decreased these effects of RasGRP3 in glioma cells. In summary, RasGRP3 acts as an important integrating signaling protein of the DAG and Ras signaling pathways and actin polymerization and represents an important therapeutic target in GBM. PMID:25682201

Economic Analysis of First-Line Treatment with Cetuximab or Panitumumab for RAS Wild-Type Metastatic Colorectal Cancer in England.

Science.gov (United States)

Tikhonova, Irina A; Huxley, Nicola; Snowsill, Tristan; Crathorne, Louise; Varley-Campbell, Jo; Napier, Mark; Hoyle, Martin

2018-03-01

Combination therapies with cetuximab (Erbitux ® ; Merck Serono UK Ltd) and panitumumab (Vectibix ® ; Amgen UK Ltd) are shown to be less effective in adults with metastatic colorectal cancer who have mutations in exons 2, 3 and 4 of KRAS and NRAS oncogenes from the rat sarcoma (RAS) family. The objective of the study was to estimate the cost effectiveness of these drugs in patients with previously untreated RAS wild-type (i.e. non-mutated) metastatic colorectal cancer, not eligible for liver resection at baseline, from the UK National Health Service and Personal Social Services perspective. We constructed a partitioned survival model to evaluate the long-term costs and benefits of cetuximab and panitumumab combined with either FOLFOX (folinic acid, fluorouracil and oxaliplatin) or FOLFIRI (folinic acid, fluorouracil and irinotecan) vs. FOLFOX or FOLFIRI alone. The economic analysis was based on three randomised controlled trials. Costs and quality-adjusted life-years were discounted at 3.5% per annum. Based on the evidence available, both drugs fulfil the National Institute for Health and Care Excellence's end-of-life criteria. In the analysis, assuming discount prices for the drugs from patient access schemes agreed by the drug manufacturers with the Department of Health, predicted mean incremental cost-effectiveness ratios for cetuximab + FOLFOX, panitumumab + FOLFOX and cetuximab + FOLFIRI compared with chemotherapy alone appeared cost-effective at the National Institute for Health and Care Excellence's threshold of £50,000 per quality-adjusted life-year gained, applicable to end-of-life treatments. Cetuximab and panitumumab were recommended by the National Institute for Health and Care Excellence for patients with previously untreated RAS wild-type metastatic colorectal cancer, not eligible for liver resection at baseline, for use within the National Health Service in England. Both treatments are available via the UK Cancer Drugs Fund.

Aberrantly methylated genes in human papillary thyroid cancer and their association with BRAF/RAS mutation.

Directory of Open Access Journals (Sweden)

Yasuko eKikuchi

2013-12-01

Full Text Available Cancer arises through accumulation of epigenetic and genetic alteration. Aberrant promoter methylation is a common epigenetic mechanism of gene silencing in cancer cells. We here performed genome-wide analysis of DNA methylation of promoter regions by Infinium HumanMethylation27 BeadChip, using 14 clinical papillary thyroid cancer samples and 10 normal thyroid samples. Among the 14 papillary cancer cases, 11 showed frequent aberrant methylation, but the other three cases showed no aberrant methylation at all. Distribution of the hypermethylation among cancer samples was non-random, which implied existence of a subset of preferentially methylated papillary thyroid cancer. Among 25 frequently methylated genes, methylation status of six genes (HIST1H3J, POU4F2, SHOX2, PHKG2, TLX3, HOXA7 was validated quantitatively by pyrosequencing. Epigenetic silencing of these genes in methylated papillary thyroid cancer cell lines was confirmed by gene re-expression following treatment with 5-aza-2'-deoxycytidine and trichostatin A, and detected by real-time RT-PCR. Methylation of these six genes was validated by analysis of additional 20 papillary thyroid cancer and 10 normal samples. Among the 34 cancer samples in total, 26 cancer samples with preferential methylation were significantly associated with mutation of BRAF/RAS oncogene (P=0.04, Fisher’s exact test. Thus we identified new genes with frequent epigenetic hypermethylation in papillary thyroid cancer, two subsets of either preferentially methylated or hardly methylated papillary thyroid cancer, with a concomitant occurrence of oncogene mutation and gene methylation. These hypermethylated genes may constitute potential biomarkers for papillary thyroid cancer.

Trabecular meshwork ECM remodeling in glaucoma: could RAS be a target?

Science.gov (United States)

Agarwal, Puneet; Agarwal, Renu

2018-06-14

Disturbances of extracellular matrix (ECM) homeostasis in trabecular meshwork (TM) cause increased aqueous outflow resistance leading to elevated intraocular pressure (IOP) in glaucomatous eyes. Therefore, restoration of ECM homeostasis is a rational approach to prevent disease progression. Since renin-angiotensin system (RAS) inhibition positively alters ECM homeostasis in cardiovascular pathologies involving pressure and volume overload, it is likely that RAS inhibitors reduce IOP primarily by restoring ECM homeostasis. Areas covered: Current evidence showing the presence of RAS components in ocular tissue and its role in regulating aqueous humor dynamics is briefly summarized. The role of RAS in ECM remodeling is discussed both in terms of its effects on ECM synthesis and its breakdown. The mechanisms of ECM remodeling involving interactions of RAS with transforming growth factor-β, Wnt/β-catenin signaling, bone morphogenic proteins, connective tissue growth factor, and matrix metalloproteinases in ocular tissue are discussed. Expert opinion: Current literature strongly indicates a significant role of RAS in ECM remodeling in TM of hypertensive eyes. Hence, IOP-lowering effect of RAS inhibitors may primarily be attributed to restoration of ECM homeostasis in aqueous outflow pathways rather than its vascular effects. However, the mechanistic targets for RAS inhibitors have much wider distribution and consequences, which remain relatively unexplored in TM.

Transformation by oncogenic RAS sensitizes human colon cells to TRAIL-induced apoptosis by up-regulating death receptor 4 and death receptor 5 through a MEK-dependent pathway

Czech Academy of Sciences Publication Activity Database

Drosopoulos, K.G.; Roberts, M.L.; Čermák, Lukáš; Sasazuki, T.; Shirasawa, S.; Anděra, Ladislav; Pintzas, A.

2005-01-01

Roč. 280, č. 24 (2005), s. 22856-22867 ISSN 0021-9258 R&D Projects: GA AV ČR(CZ) KSK5020115; GA AV ČR(CZ) KJB5052407 Keywords : TRAIL * Ras * apoptosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.854, year: 2005

Non-covalent interactions of the carcinogen (+)-anti-BPDE with exon 1 of the human K-ras proto-oncogene

Science.gov (United States)

Rodriguez, Jorge H.; Deligkaris, Christos

2013-03-01

Investigating the complementary, but different, effects of physical (non-covalent) and chemical (covalent) mutagen-DNA and carcinogen-DNA interactions is important for understanding possible mechanisms of development and prevention of mutagenesis and carcinogenesis. A highly mutagenic and carcinogenic metabolite of the polycyclic aromatic hydrocarbon benzo[ α]pyrene, namely (+)-anti-BPDE, is known to undergo both physical and chemical complexation with DNA. The major covalent adduct, a promutagenic, is known to be an external (+)-trans-anti-BPDE-N2-dGuanosine configuration whose origins are not fully understood. Thus, it is desirable to study the mechanisms of external non-covalent BPDE-DNA binding and their possible relationships to external covalent trans adduct formation. We present a detailed codon-by-codon computational study of the non-covalent interactions of (+)-anti-BPDE with DNA which explains and correctly predicts preferential (+)-anti-BPDE binding at minor groove guanosines. Due to its relevance to carcinogenesis, the interaction of (+)-anti-BPDE with exon 1 of the human K-ras gene has been studied in detail. Present address: Department of Physics, Drury University

Spectrum of K ras mutations in Pakistani colorectal cancer patients

International Nuclear Information System (INIS)

Murtaza, B.N.; Bibi, A.; Rashid, M.U.; Khan, Y.I.; Chaudri, M.S.; Shakoori, A.R.

2013-01-01

The incidence of colorectal cancer (CRC) is increasing daily worldwide. Although different aspects of CRC have been studied in other parts of the world, relatively little or almost no information is available in Pakistan about different aspects of this disease at the molecular level. The present study was aimed at determining the frequency and prevalence of K ras gene mutations in Pakistani CRC patients. Tissue and blood samples of 150 CRC patients (64% male and 36% female) were used for PCR amplification of K ras and detection of mutations by denaturing gradient gel electrophoresis, restriction fragment length polymorphism analysis, and nucleotide sequencing. The K ras mutation frequency was found to be 13%, and the most prevalent mutations were found at codons 12 and 13. A novel mutation was also found at codon 31. The dominant mutation observed was a G to A transition. Female patients were more susceptible to K ras mutations, and these mutations were predominant in patients with a nonmetastatic stage of CRC. No significant differences in the prevalence of K ras mutations were observed for patient age, gender, or tumor type. It can be inferred from this study that Pakistani CRC patients have a lower frequency of K ras mutations compared to those observed in other parts of the world, and that K ras mutations seemed to be significantly associated with female patients

Spectrum of K ras mutations in Pakistani colorectal cancer patients

Energy Technology Data Exchange (ETDEWEB)

Murtaza, B.N.; Bibi, A. [School of Biological Sciences, University of the Punjab, Quaid-i-Azam Campus, Lahore (Pakistan); Rashid, M.U.; Khan, Y.I. [Shaukat Khanum Memorial Cancer Hospital and Research Centre, Johar Town, Lahore (Pakistan); Chaudri, M.S. [Services Institute of Medical Sciences, Lahore (Pakistan); Shakoori, A.R. [School of Biological Sciences, University of the Punjab, Quaid-i-Azam Campus, Lahore (Pakistan)

2013-11-29

The incidence of colorectal cancer (CRC) is increasing daily worldwide. Although different aspects of CRC have been studied in other parts of the world, relatively little or almost no information is available in Pakistan about different aspects of this disease at the molecular level. The present study was aimed at determining the frequency and prevalence of K ras gene mutations in Pakistani CRC patients. Tissue and blood samples of 150 CRC patients (64% male and 36% female) were used for PCR amplification of K ras and detection of mutations by denaturing gradient gel electrophoresis, restriction fragment length polymorphism analysis, and nucleotide sequencing. The K ras mutation frequency was found to be 13%, and the most prevalent mutations were found at codons 12 and 13. A novel mutation was also found at codon 31. The dominant mutation observed was a G to A transition. Female patients were more susceptible to K ras mutations, and these mutations were predominant in patients with a nonmetastatic stage of CRC. No significant differences in the prevalence of K ras mutations were observed for patient age, gender, or tumor type. It can be inferred from this study that Pakistani CRC patients have a lower frequency of K ras mutations compared to those observed in other parts of the world, and that K ras mutations seemed to be significantly associated with female patients.

Ras Activity Oscillates in the Mouse Suprachiasmatic Nucleus and Modulates Circadian Clock Dynamics.

Science.gov (United States)

Serchov, Tsvetan; Jilg, Antje; Wolf, Christian T; Radtke, Ina; Stehle, Jörg H; Heumann, Rolf

2016-04-01

Circadian rhythms, generated in the mouse suprachiasmatic nucleus (SCN), are synchronized to the environmental day-night changes by photic input. The activation of the extracellular signal-regulated kinases 1 and 2 (ERK1,2) and cAMP response element-binding protein (CREB)-mediated transcription play a critical role in this photoentrainment. The small GTPase Ras is one of the major upstream regulators of the ERK1,2/CREB pathway. In contrast to the well-described role of Ras in structural and functional synaptic plasticity in the adult mouse brain, the physiological regulation of Ras by photic sensory input is yet unknown. Here, we describe for the first time a circadian rhythm of Ras activity in the mouse SCN. Using synRas transgenic mice, expressing constitutively activated V12-Ha-Ras selectively in neurons, we demonstrate that enhanced Ras activation causes shortening of the circadian period length. We found upregulated expression and decreased inhibitory phosphorylation of the circadian period length modulator, glycogen synthase kinase-3 beta (GSK3β), in the SCN of synRas mice. Conversely, downregulation of Ras activity by blocking its function with an antibody in oscillating cell cultures reduced protein levels and increased phosphorylation of GSK3β and lengthened the period of BMAL1 promoter-driven luciferase activity. Furthermore, enhanced Ras activity in synRas mice resulted in a potentiation of light-induced phase delays at early subjective night, and increased photic induction of pERK1,2/pCREB and c-Fos. In contrast, at late subjective night, photic activation of Ras/ERK1,2/CREB in synRas mice was not sufficient to stimulate c-Fos protein expression and phase advance the clock. Taken together, our results demonstrate that Ras activity fine tunes the period length and modulates photoentrainment of the circadian clock.

Genetic and Molecular Analysis of Suppressors of Ras Mutations

National Research Council Canada - National Science Library

Eastburn, Dennis

2000-01-01

.... The study of Caenorhabditis elegans and other model systems has demonstrated that Ras is part of a conserved Ras/MAPK signaling pathway involved in many aspects of development and cell regulation. The C...

Quantitative Assays for RAS Pathway Proteins and Phosphorylation States

Science.gov (United States)

The NCI CPTAC program is applying its expertise in quantitative proteomics to develop assays for RAS pathway proteins. Targets include key phosphopeptides that should increase our understanding of how the RAS pathway is regulated.

Interaction of CtBP with adenovirus E1A suppresses immortalization of primary epithelial cells and enhances virus replication during productive infection

Energy Technology Data Exchange (ETDEWEB)

Subramanian, T.; Zhao, Ling-jun; Chinnadurai, G., E-mail: chinnag@slu.edu

2013-09-01

Adenovirus E1A induces cell proliferation, oncogenic transformation and promotes viral replication through interaction with p300/CBP, TRRAP/p400 multi-protein complex and the retinoblastoma (pRb) family proteins through distinct domains in the E1A N-terminal region. The C-terminal region of E1A suppresses E1A/Ras co-transformation and interacts with FOXK1/K2, DYRK1A/1B/HAN11 and CtBP1/2 (CtBP) protein complexes. To specifically dissect the role of CtBP interaction with E1A, we engineered a mutation (DL→AS) within the CtBP-binding motif, PLDLS, and investigated the effect of the mutation on immortalization and Ras cooperative transformation of primary cells and viral replication. Our results suggest that CtBP–E1A interaction suppresses immortalization and Ras co-operative transformation of primary rodent epithelial cells without significantly influencing the tumorigenic activities of transformed cells in immunodeficient and immunocompetent animals. During productive infection, CtBP–E1A interaction enhances viral replication in human cells. Between the two CtBP family proteins, CtBP2 appears to restrict viral replication more than CtBP1 in human cells. - Highlights: • Adenovirus E1A C-terminal region suppresses E1A/Ras co-transformation. • This E1A region binds with FOXK, DYRK1/HAN11 and CtBP cellular protein complexes. • We found that E1A–CtBP interaction suppresses immortalization and transformation. • The interaction enhances viral replication in human cells.

Modelling Cooperative Tumorigenesis in Drosophila

Science.gov (United States)

2018-01-01

The development of human metastatic cancer is a multistep process, involving the acquisition of several genetic mutations, tumour heterogeneity, and interactions with the surrounding microenvironment. Due to the complexity of cancer development in mammals, simpler model organisms, such as the vinegar fly, Drosophila melanogaster, are being utilized to provide novel insights into the molecular mechanisms involved. In this review, we highlight recent advances in modelling tumorigenesis using the Drosophila model, focusing on the cooperation of oncogenes or tumour suppressors, and the interaction of mutant cells with the surrounding tissue in epithelial tumour initiation and progression. PMID:29693007

Modelling Cooperative Tumorigenesis in Drosophila

Directory of Open Access Journals (Sweden)

Helena E. Richardson

2018-01-01

Full Text Available The development of human metastatic cancer is a multistep process, involving the acquisition of several genetic mutations, tumour heterogeneity, and interactions with the surrounding microenvironment. Due to the complexity of cancer development in mammals, simpler model organisms, such as the vinegar fly, Drosophila melanogaster, are being utilized to provide novel insights into the molecular mechanisms involved. In this review, we highlight recent advances in modelling tumorigenesis using the Drosophila model, focusing on the cooperation of oncogenes or tumour suppressors, and the interaction of mutant cells with the surrounding tissue in epithelial tumour initiation and progression.

Phenotypic Screening Identifies Protein Synthesis Inhibitors as H-Ras-Nanocluster-Increasing Tumor Growth Inducers.

Science.gov (United States)

Najumudeen, Arafath K; Posada, Itziar M D; Lectez, Benoit; Zhou, Yong; Landor, Sebastian K-J; Fallarero, Adyary; Vuorela, Pia; Hancock, John; Abankwa, Daniel

2015-12-15

Ras isoforms H-, N-, and K-ras are each mutated in specific cancer types at varying frequencies and have different activities in cell fate control. On the plasma membrane, Ras proteins are laterally segregated into isoform-specific nanoscale signaling hubs, termed nanoclusters. As Ras nanoclusters are required for Ras signaling, chemical modulators of nanoclusters represent ideal candidates for the specific modulation of Ras activity in cancer drug development. We therefore conducted a chemical screen with commercial and in-house natural product libraries using a cell-based H-ras-nanoclustering FRET assay. Next to established Ras inhibitors, such as a statin and farnesyl-transferase inhibitor, we surprisingly identified five protein synthesis inhibitors as positive regulators. Using commonly employed cycloheximide as a representative compound, we show that protein synthesis inhibition increased nanoclustering and effector recruitment specifically of active H-ras but not of K-ras. Consistent with these data, cycloheximide treatment activated both Erk and Akt kinases and specifically promoted H-rasG12V-induced, but not K-rasG12V-induced, PC12 cell differentiation. Intriguingly, cycloheximide increased the number of mammospheres, which are enriched for cancer stem cells. Depletion of H-ras in combination with cycloheximide significantly reduced mammosphere formation, suggesting an exquisite synthetic lethality. The potential of cycloheximide to promote tumor cell growth was also reflected in its ability to increase breast cancer cell tumors grown in ovo. These results illustrate the possibility of identifying Ras-isoform-specific modulators using nanocluster-directed screening. They also suggest an unexpected feedback from protein synthesis inhibition to Ras signaling, which might present a vulnerability in certain tumor cell types.

The ras1 function of Schizosaccharomyces pombe mediates pheromone-induced transcription

DEFF Research Database (Denmark)

Nielsen, O; Davey, William John; Egel, R

1992-01-01

Loss of ras1+ function renders fission yeast cells unable to undergo morphological changes in response to mating pheromones, whereas cells carrying activated mutations in ras1 are hyper-responsive. This has led to the suggestion that the ras1 gene product plays a role in mating pheromone signal...

Orphan nuclear receptor TLX functions as a potent suppressor of oncogene-induced senescence in prostate cancer via its transcriptional co-regulation of the CDKN1A (p21(WAF1) (/) (CIP1) ) and SIRT1 genes.

Science.gov (United States)

Wu, Dinglan; Yu, Shan; Jia, Lin; Zou, Chang; Xu, Zhenyu; Xiao, Lijia; Wong, Kam-Bo; Ng, Chi-Fai; Chan, Franky L

2015-05-01

Oncogene-induced senescence is an important tumour-suppressing mechanism to prevent both premalignant transformation and cancer progression. Overcoming this process is a critical step in early cancer development. The druggable orphan nuclear receptor TLX (NR2E1) is characterized as an important regulator of neural stem cells and is also implicated in the development of some brain tumours. However, its exact functional roles in cancer growth regulation still remain unclear. Here we report that TLX can act as a promoter of tumourigenesis in prostate cancer by suppressing oncogene-induced senescence. We determined that TLX exhibited an increased expression in high-grade prostate cancer tissues and many prostate cancer cell lines. Functional studies revealed that TLX could perform an oncogenic function in prostate cancer cells, as its knockdown triggered cellular senescence and cell growth arrest in vitro and in vivo, whereas its over-expression promoted the malignant growth of prostate cancer cells. Furthermore, enhancement of TLX activity, by either ectopic expression or ligand stimulation, could potently prevent doxorubicin-induced senescence in prostate cancer cells and also allow prostatic epithelial cells to escape oncogene-induced senescence induced either by activated oncogene H-Ras(G12V) or knockdown of tumour suppressor PTEN, via a mechanism of direct but differential transcriptional regulation of two senescence-associated genes, repression of CDKN1A and transactivation of SIRT1. Together, our present study shows, for the first time, that TLX may play an important role in prostate carcinogenesis through its suppression of oncogene-induced senescence, and also suggests that targeting the senescence-regulatory TLX is of potential therapeutic significance in prostate cancer. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

RAS signalling in energy metabolism and rare human diseases.

Science.gov (United States)

Dard, L; Bellance, N; Lacombe, D; Rossignol, R

2018-05-08

The RAS pathway is a highly conserved cascade of protein-protein interactions and phosphorylation that is at the heart of signalling networks that govern proliferation, differentiation and cell survival. Recent findings indicate that the RAS pathway plays a role in the regulation of energy metabolism via the control of mitochondrial form and function but little is known on the participation of this effect in RAS-related rare human genetic diseases. Germline mutations that hyperactivate the RAS pathway have been discovered and linked to human developmental disorders that are known as RASopathies. Individuals with RASopathies, which are estimated to affect approximately 1/1000 human birth, share many overlapping characteristics, including cardiac malformations, short stature, neurocognitive impairment, craniofacial dysmorphy, cutaneous, musculoskeletal, and ocular abnormalities, hypotonia and a predisposition to developing cancer. Since the identification of the first RASopathy, type 1 neurofibromatosis (NF1), which is caused by the inactivation of neurofibromin 1, several other syndromes have been associated with mutations in the core components of the RAS-MAPK pathway. These syndromes include Noonan syndrome (NS), Noonan syndrome with multiple lentigines (NSML), which was formerly called LEOPARD syndrome, Costello syndrome (CS), cardio-facio-cutaneous syndrome (CFC), Legius syndrome (LS) and capillary malformation-arteriovenous malformation syndrome (CM-AVM). Here, we review current knowledge about the bioenergetics of the RASopathies and discuss the molecular control of energy homeostasis and mitochondrial physiology by the RAS pathway. Copyright © 2018 Elsevier B.V. All rights reserved.

Coexistence of K-ras mutations and HPV infection in colon cancer

Directory of Open Access Journals (Sweden)

Tezol Ayda

2006-05-01

Full Text Available Abstract Background Activation of the ras genes or association with human papillomavirus infection have been extensively studied in colorectal cancer. However, the correlation between K-ras mutations and HPV in colorectal cancer has not been investigated yet. In this study we aimed to investigate the presence of K-ras mutations and their correlation with HPV infection in colon cancer. Methods K-ras mutations were analyzed by a mutagenic PCR assay and digestion with specific restriction enzymes to distinguish the wild-type and mutant codons. HPV infection was analyzed by PCR amplification and hybridization with specific probes by Southern blotting. Stattistical analyses were performed by the chi-square and Fisher's exact tests Results HPV gene fragments were detected in 43 tumors and 17 normal tissue samples. HPV 18 was the prevalent type in the tumor tissue. A mutation at codon 12 of the K-ras gene was present in 31 patients. 56% of the HPV-positive tumors also harbored a K-ras mutation. Codon 13 mutations were not observed. These data indicate that infection with high risk HPV types and mutational activation of the K-ras gene are frequent events in colorectal carcinogenesis. Conclusion Our findings suggest that mutational activation of the K-ras gene is a common event in colon carcinogenesis and that HPV infection may represent an important factor in the development of the premalignant lesions leading to the neoplastic phenotype.

[Carcinogenesis and its mechanism of mutant-type[12Asp]K-ras4B gene].

Science.gov (United States)

Gui, Li-ming; Wei, Li-hui; Zhang, Ying-mei; Wang, Jian-liu; Wang, Ying; Chen, Ying; Ma, Da-long

2002-01-01

Ras gene plays an important role in the extra- and intra-cellular signal transduction pathway. It mediates series cascade reactions, and eventually actives transcriptional factors in nucleus. It is unknown on the mechanism of carcinogenesis of Ras gene in endometrial carcinoma, though K-ras mutant is very common in endometrial atypical hyperplasia and carcinoma. On basis of discovering the mutation in 12th codon of K-ras in endometrial carcinoma cell line, HEC-1A, we explored the carcinogenesis and molecular mechanism of mutant-type [12Asp] K-ras4B gene. (1) Full-length [12Asp]K-ras4B cDNA was amplified with RT-PCR, then inserted into pcDI eukaryotic expressive vector. (2) Morphological change, growth kinetics in vitro and tumorigencity in nude mice in vivo after-before transfection were observed. (3) To test the cell growth kinetics by methyl thiazolium tetrazolium (MTT) and [3H]thymidine incorporation method. (1) The authors have successfully constructed eukaryotic expression plasmid pcDI-[12Asp] K-ras4B; (2) To confirm that [12Asp] K-ras4B mutant can trigger the neoplastic transformation of NIH3T3 cells by test in vitro and in vivo. (3) After pMCV-RasN17 plasmid, a Ras mutant were transfected into pcDI-[12Asp] K-ras4B cells, the growth of this cell were restrained significantly in comparison with control group. (4) These findings indicate the expression of RafS621A resulted in remarkable inhibition in proliferation of pcDI-[12Asp]K-ras4B cell (P ras4B cell growth (P ras4B gene alone is able to cause neoplastic transformation in NIH3T3 cells in vitro and in vivo. (2) [12Asp]K-ras4B-induced NIH3T3 cells neoplastic transformation required Raf signaling pathway.

RasC is required for optimal activation of adenylyl cyclase and Akt/PKB during aggregation.

Science.gov (United States)

Lim, C J; Spiegelman, G B; Weeks, G

2001-08-15

Disruption of Dictyostelium rasC, encoding a Ras subfamily protein, generated cells incapable of aggregation. While rasC expression is enriched in a cell type-specific manner during post-aggregative development, the defect in rasC(-) cells is restricted to aggregation and fully corrected by application of exogenous cAMP pulses. cAMP is not produced in rasC(-) cells stimulated by 2'-deoxy-cAMP, but is produced in response to GTPgammaS in cell lysates, indicating that G-protein-coupled cAMP receptor activation of adenylyl cyclase is regulated by RasC. However, cAMP-induced ERK2 phosphorylation is unaffected in rasC(-) cells, indicating that RasC is not an upstream activator of the mitogen-activated protein kinase required for cAMP relay. rasC(-) cells also exhibit reduced chemotaxis to cAMP during early development and delayed response to periodic cAMP stimuli produced by wild-type cells in chimeric mixtures. Furthermore, cAMP-induced Akt/PKB phosphorylation through a phosphatidylinositide 3-kinase (PI3K)-dependent pathway is dramatically reduced in rasC(-) cells, suggesting that G-protein-coupled serpentine receptor activation of PI3K is regulated by RasC. Cells lacking the RasGEF, AleA, exhibit similar defects as rasC(-) cells, suggesting that AleA may activate RasC.

Promoter methylation of RASSF1A and DAPK and mutations of K-ras, p53, and EGFR in lung tumors from smokers and never-smokers

International Nuclear Information System (INIS)

Liu, Yang; Gao, Weimin; Siegfried, Jill M; Weissfeld, Joel L; Luketich, James D; Keohavong, Phouthone

2007-01-01

Epidemiological studies indicate that some characteristics of lung cancer among never-smokers significantly differ from those of smokers. Aberrant promoter methylation and mutations in some oncogenes and tumor suppressor genes are frequent in lung tumors from smokers but rare in those from never-smokers. In this study, we analyzed promoter methylation in the ras-association domain isoform A (RASSF1A) and the death-associated protein kinase (DAPK) genes in lung tumors from patients with primarily non-small cell lung cancer (NSCLC) from the Western Pennsylvania region. We compare the results with the smoking status of the patients and the mutation status of the K-ras, p53, and EGFR genes determined previously on these same lung tumors. Promoter methylation of the RASSF1A and DAPK genes was analyzed by using a modified two-stage methylation-specific PCR. Data on mutations of K-ras, p53, and EGFR were obtained from our previous studies. The RASSF1A gene promoter methylation was found in tumors from 46.7% (57/122) of the patients and was not significantly different between smokers and never-smokers, but was associated significantly in multiple variable analysis with tumor histology (p = 0.031) and marginally with tumor stage (p = 0.063). The DAPK gene promoter methylation frequency in these tumors was 32.8% (40/122) and did not differ according to the patients' smoking status, tumor histology, or tumor stage. Multivariate analysis adjusted for age, gender, smoking status, tumor histology and stage showed that the frequency of promoter methylation of the RASSF1A or DAPK genes did not correlate with the frequency of mutations of the K-ras, p53, and EGFR gene. Our results showed that RASSF1A and DAPK genes' promoter methylation occurred frequently in lung tumors, although the prevalence of this alteration in these genes was not associated with the smoking status of the patients or the occurrence of mutations in the K-ras, p53 and EGFR genes, suggesting each of

Ras activation by SOS

DEFF Research Database (Denmark)

Iversen, Lars; Tu, Hsiung-Lin; Lin, Wan-Chen

2014-01-01

Activation of the small guanosine triphosphatase H-Ras by the exchange factor Son of Sevenless (SOS) is an important hub for signal transduction. Multiple layers of regulation, through protein and membrane interactions, govern activity of SOS. We characterized the specific activity of individual ...

A Network-Based Model of Oncogenic Collaboration for Prediction of Drug Sensitivity

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Ted G Laderas

2015-12-01

Full Text Available Tumorigenesis is a multi-step process, involving the acquisition of multiple oncogenic mutations that transform cells, resulting in systemic dysregulation that enables proliferation, among other cancer hallmarks. High throughput omics techniques are used in precision medicine, allowing identification of these mutations with the goal of identifying treatments that target them. However, the multiplicity of oncogenes required for transformation, known as oncogenic collaboration, makes assigning effective treatments difficult. Motivated by this observation, we propose a new type of oncogenic collaboration where mutations in genes that interact with an oncogene may contribute to its dysregulation, a new genomic feature that we term surrogate oncogenes. By mapping mutations to a protein/protein interaction network, we can determine significance of the observed distribution using permutation-based methods. For a panel of 38 breast cancer cell lines, we identified significant surrogate oncogenes in oncogenes such as BRCA1 and ESR1. In addition, using Random Forest Classifiers, we show that these significant surrogate oncogenes predict drug sensitivity for 74 drugs in the breast cancer cell lines with a mean error rate of 30.9%. Additionally, we show that surrogate oncogenes are predictive of survival in patients. The surrogate oncogene framework incorporates unique or rare mutations on an individual level. Our model has the potential for integrating patient-unique mutations in predicting drug-sensitivity, suggesting a potential new direction in precision medicine, as well as a new approach for drug development. Additionally, we show the prevalence of significant surrogate oncogenes in multiple cancers within the Cancer Genome Atlas, suggesting that surrogate oncogenes may be a useful genomic feature for guiding pancancer analyses and assigning therapies across many tissue types.

Genomewide Search of Oncogenic Pathways Cooperating With ETS Fusions in Prostate Cancer

Science.gov (United States)

2014-09-01

Arrows in T-ETV1 pictures indicate inflammation in T-ETV1 knock-in males ([left] slight inflammation in the lateral lobe of a young knock-in male...drives anabolic metabolism and tumorigenesis (Ward and Thompson 2012). We propose that TMPRSS2-ETV1 cooperates with Pten loss by further enhancing

RasGRP1 confers the phorbol ester-sensitive phenotype to EL4 lymphoma cells.

Science.gov (United States)

Han, Shujie; Knoepp, Stewart M; Hallman, Mark A; Meier, Kathryn E

2007-01-01

The murine EL4 lymphoma cell line exists in variants that are either sensitive or resistant to the tumor promoter phorbol 12-myristate 13-acetate (PMA). In sensitive EL4 cells, PMA causes robust Erk mitogen-activated protein kinase activation that results in growth arrest. In resistant cells, PMA induces minimal Erk activation, without growth arrest. PMA stimulates IL-2 production in sensitive, but not resistant, cells. The role of RasGRP1, a PMA-activated guanine nucleotide exchange factor for Ras, in EL4 phenotype was examined. Endogenous RasGRP1 protein is expressed at much higher levels in sensitive than in resistant cells. PMA-induced Ras activation is observed in sensitive cells but not in resistant cells lacking Ras-GRP1. PMA induces down-regulation of RasGRP1 protein in sensitive cells but increases RasGRP1 in resistant cells. Transfection of RasGRP1 into resistant cells enhances PMA-induced Erk activation. In the reverse experiment, introduction of small interfering RNA (siRNA) for RasGRP1 suppresses PMA-induced Ras and Erk activations in sensitive cells. Sensitive cells incubated with siRNA for RasGRP1 exhibit the PMA-resistant phenotype, in that they are able to proliferate in the presence of PMA and do not secrete IL-2 when stimulated with PMA. These studies indicate that the PMA-sensitive phenotype, as previously defined for the EL4 cell line, is conferred by endogenous expression of RasGRP1 protein.

Changes in microbial water quality in RAS following altered feed loading

DEFF Research Database (Denmark)

Rojas-Tirado, Paula Andrea; Pedersen, Per Bovbjerg; Vadstein, Olav

2018-01-01

and inorganic nutrients available for microbial growth in RAS. How these nutrient inputs affect and regulate bacteria in RAS water is, however, unclear. To investigate this relationship and the associated water quality dynamics, the effects of altered feed loading on microbial water quality in RAS was studied....... The study included six independent, identical pilot-scale RAS, each with a total volume of 1.7 m3 (make-up water: 80 L/day) stocked with juvenile rainbow trout (Oncorhynchus mykiss). All systems had been operating with constant and identical feed loading of 3.13 kg feed/m3 make-up water for a period......Intensive recirculating aquaculture systems (RAS) with its hyper-eutrophic water offer ideal conditions for bacterial growth, abundance and activity, potentially affecting fish and system performance. Feed composition and feed loading in particular will have significant impact on organic...

Identification of Differentially Expressed K-Ras Transcript Variants in Patients With Leiomyoma.

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Zolfaghari, Nooshin; Shahbazi, Shirin; Torfeh, Mahnaz; Khorasani, Maryam; Hashemi, Mehrdad; Mahdian, Reza

2017-10-01

Molecular studies have demonstrated a wide range of gene expression variations in uterine leiomyoma. The rat sarcoma virus/rapidly accelerated fibrosarcoma/mitogen-activated protein kinase (RAS/RAF/MAPK) is the crucial cellular pathway in transmitting external signals into nucleus. Deregulation of this pathway contributes to excessive cell proliferation and tumorigenesis. The present study aims to investigate the expression profile of the K-Ras transcripts in tissue samples from patients with leiomyoma. The patients were leiomyoma cases who had no mutation in mediator complex subunit 12 ( MED12) gene. A quantitative approach has been applied to determine the difference in the expression of the 2 main K-Ras messenger RNA (mRNA) variants. The comparison between gene expression levels in leiomyoma and normal myometrium group was performed using relative expression software tool. The expression of K-Ras4B gene was upregulated in leiomyoma group ( P = .016), suggesting the involvement of K-Ras4B in the disease pathogenesis. Pairwise comparison of the K-Ras4B expression between each leiomyoma tissue and its matched adjacent normal myometrium revealed gene upregulation in 68% of the cases. The expression of K-Ras4A mRNA was relatively upregulated in leiomyoma group ( P = .030). In addition, the mean expression of K-Ras4A gene in leiomyoma tissues relative to normal samples was 4.475 (95% confidence interval: 0.10-20.42; standard error: 0.53-12.67). In total, 58% of the cases showed more than 2-fold increase in K-Ras4A gene expression. Our results demonstrated increased expression of both K-Ras mRNA splicing variants in leiomyoma tissue. However, the ultimate result of KRAS expression on leiomyoma development depends on the overall KRAS isoform balance and, consequently, on activated signaling pathways.

Oncogene mutational profile in nasopharyngeal carcinoma

Directory of Open Access Journals (Sweden)

Zhang ZC

2014-03-01

Full Text Available Zi-Chen Zhang,1,* Sha Fu,1,* Fang Wang,1 Hai-Yun Wang,1 Yi-Xin Zeng,2 Jian-Yong Shao11Department of Molecular Diagnostics, 2Department of Experimental Research, Sun Yat-sen University Cancer Center, State Key Laboratory of Oncology in South China, Collaborative Innovation Center of Cancer Medicine, Guangzhou, People's Republic of China *These authors contributed equally to this work Abstract: Nasopharyngeal carcinoma (NPC is a common tumor in Southern China, but the oncogene mutational status of NPC patients has not been clarified. Using time-of-flight mass spectrometry, 238 mutation hotspots in 19 oncogenes were examined in 123 NPC patients. The relationships between mutational status and clinical data were assessed with a χ2 or Fisher's exact test. Survival analysis was performed using the Kaplan–Meier method with the log-rank test. In 123 patients, 21 (17.1% NPC tumors were positive for mutations in eight oncogenes: six patients had PIK3CA mutations (4.9%, five NRAS mutations (4.1%, four KIT mutations (3.3%, two PDGFRA mutations (1.6%, two ABL mutations (1.6%, and one with simultaneous mutations in HRAS, EGFR, and BRAF (1%. Patients with mutations were more likely to relapse or develop metastasis than those with wild-type alleles (P=0.019. No differences or correlations were found in other clinical characteristics or in patient survival. No mutations were detected in oncogenes AKT1, AKT2, CDK, ERBB2, FGFR1, FGFR3, FLT3, JAK2, KRAS, MET, and RET. These results demonstrate an association between NPC and mutations in NRAS, KIT, PIK3CA, PDGFRA, and ABL, which are associated with patient relapse and metastasis. Keywords: NPC, oncogene, mutation

Genetic Validation of Cell Proliferation via Ras-Independent Activation of the Raf/Mek/Erk Pathway.

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Lechuga, Carmen G; Simón-Carrasco, Lucía; Jacob, Harrys K C; Drosten, Matthias

2017-01-01

Signaling transmitted by the Ras family of small GTPases (H-, N-, and K-Ras) is essential for proliferation of mouse embryonic fibroblasts (MEFs). However, constitutive activation of the downstream Raf/Mek/Erk pathway can bypass the requirement for Ras proteins and allow cells to proliferate in the absence of the three Ras isoforms. Here we describe a protocol for a colony formation assay that permits evaluating the role of candidate proteins that are positive or negative regulators of cell proliferation mediated via Ras-independent Raf/Mek/Erk pathway activation. K-Ras lox (H-Ras -/- , N-Ras -/- , K-Ras lox/lox , RERT ert/ert ) MEFs are infected with retro- or lentiviral vectors expressing wild-type or constitutively activated candidate cDNAs, shRNAs, or sgRNAs in combination with Cas9 to ascertain the possibility of candidate proteins to function either as an activator or inhibitor of Ras-independent Raf/Mek/Erk activation. These cells are then seeded in the absence or presence of 4-Hydroxytamoxifen (4-OHT), which activates the resident CreERT2 alleles resulting in elimination of the conditional K-Ras alleles and ultimately generating Rasless cells. Colony formation in the presence of 4-OHT indicates cell proliferation via Ras-independent Raf/Mek/Erk activation.

Ran is a potential therapeutic target for cancer cells with molecular changes associated with activation of the PI3K/Akt/mTORC1 and Ras/MEK/ERK pathways

Science.gov (United States)

Yuen, Hiu-Fung; Chan, Ka-Kui; Grills, Claire; Murray, James T.; Platt-Higgins, Angela; Eldin, Osama Sharaf; O’Byrne, Ken; Janne, Pasi; Fennell, Dean A.; Johnston, Patrick G.; Rudland, Philip S.; El-Tanani, Mohamed

2011-01-01

Purpose Cancer cells have been shown to be more susceptible to Ran knockdown compared to normal cells. We now investigate whether Ran is a potential therapeutic target of cancers with frequently found mutations that lead to higher Ras/MEK/ERK and PI3K/Akt/mTORC1 activities. Experimental Design Apoptosis was measured by flow cytometry (PI and Annexin V staining) and MTT assay in cancer cells grown under different conditions after knockdown of Ran.. The correlations between Ran expression and patient survival were examined in breast and lung cancers. Results Cancer cells with their PI3K/Akt/mTORC1 and Ras/MEK/ERK pathways inhibited are less susceptible to Ran silencing-induced apoptosis. KRas mutated, c-Met amplified and Pten-deleted cancer cells are also more susceptible to Ran silencing-induced apoptosis than their wild-type counterparts and this effect is reduced by inhibitors of the PI3K/Akt/mTORC1 and MEK/ERK pathways. Overexpression of Ran in clinical specimens is significantly associated with poor patient outcome in both breast and lung cancers. This association is dramatically enhanced in cancers with increased c-Met or osteopontin expression, or with oncogenic mutations of KRas or PIK3CA, all of which are mutations that potentially correlate with activation of the PI3K/Akt/mTORC1 and/or Ras/MEK/ERK pathways. Silencing Ran also results in dysregulation of nucleocytoplasmic transport of transcription factors and downregulation of Mcl-1 expression, at the transcriptional level, which are reversed by inhibitors of the PI3K/Akt/mTORC1 and MEK/ERK pathways. Conclusion Ran is a potential therapeutic target for treatment of cancers with mutations/changes of expression in protooncogenes that lead to activation of the PI3K/Akt/mTORC1 and Ras/MEK/ERK pathways. PMID:22090358

Ras GTPases Modulate Morphogenesis, Sporulation and Cellulase Gene Expression in the Cellulolytic Fungus Trichoderma reesei

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Zhang, Jiwei; Zhang, Yanmei; Zhong, Yaohua; Qu, Yinbo; Wang, Tianhong

2012-01-01

Background The model cellulolytic fungus Trichoderma reesei (teleomorph Hypocrea jecorina) is capable of responding to environmental cues to compete for nutrients in its natural saprophytic habitat despite its genome encodes fewer degradative enzymes. Efficient signalling pathways in perception and interpretation of environmental signals are indispensable in this process. Ras GTPases represent a kind of critical signal proteins involved in signal transduction and regulation of gene expression. In T. reesei the genome contains two Ras subfamily small GTPases TrRas1 and TrRas2 homologous to Ras1 and Ras2 from S. cerevisiae, but their functions remain unknown. Methodology/Principal Findings Here, we have investigated the roles of GTPases TrRas1 and TrRas2 during fungal morphogenesis and cellulase gene expression. We show that both TrRas1 and TrRas2 play important roles in some cellular processes such as polarized apical growth, hyphal branch formation, sporulation and cAMP level adjustment, while TrRas1 is more dominant in these processes. Strikingly, we find that TrRas2 is involved in modulation of cellulase gene expression. Deletion of TrRas2 results in considerably decreased transcription of cellulolytic genes upon growth on cellulose. Although the strain carrying a constitutively activated TrRas2G16V allele exhibits increased cellulase gene transcription, the cbh1 and cbh2 expression in this mutant still strictly depends on cellulose, indicating TrRas2 does not directly mediate the transmission of the cellulose signal. In addition, our data suggest that the effect of TrRas2 on cellulase gene is exerted through regulation of transcript abundance of cellulase transcription factors such as Xyr1, but the influence is independent of cAMP signalling pathway. Conclusions/Significance Together, these findings elucidate the functions for Ras signalling of T. reesei in cellular morphogenesis, especially in cellulase gene expression, which contribute to deciphering the

Reduced HRAS G12V-Driven Tumorigenesis of Cell Lines Expressing KRAS C118S.

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Lu Huang

Full Text Available In many different human cancers, one of the HRAS, NRAS, or KRAS genes in the RAS family of small GTPases acquires an oncogenic mutation that renders the encoded protein constitutively GTP-bound and thereby active, which is well established to promote tumorigenesis. In addition to oncogenic mutations, accumulating evidence suggests that the wild-type isoforms may also be activated and contribute to oncogenic RAS-driven tumorigenesis. In this regard, redox-dependent reactions with cysteine 118 (C118 have been found to promote activation of wild-type HRAS and NRAS. We sought to determine if this residue is also important for the activation of wild-type KRAS and promotion of tumorigenesis. Thus, we mutated C118 to serine (C118S in wild-type KRAS to block redox-dependent reactions at this site. We now report that this mutation reduced the level of GTP-bound KRAS and impaired RAS signaling stimulated by the growth factor EGF. With regards to tumorigenesis, we also report that oncogenic HRAS-transformed human cells in which endogenous KRAS was knocked down and replaced with KRASC118S exhibited reduced xenograft tumor growth, as did oncogenic HRAS-transformed KrasC118S/C118S murine cells in which the C118S mutation was knocked into the endogenous Kras gene. Taken together, these data suggest a role for redox-dependent activation of wild-type KRAS through C118 in oncogenic HRAS-driven tumorigenesis.

Impact of Emergent Circulating Tumor DNA RAS Mutation in Panitumumab-Treated Chemoresistant Metastatic Colorectal Cancer.

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Kim, Tae Won; Peeters, Marc; Thomas, Anne L; Gibbs, Peter; Hool, Kristina; Zhang, Jianqi; Ang, Agnes; Bach, Bruce Allen; Price, Timothy

2018-06-13

The accumulation of emergent RAS mutations during anti-epidermal growth factor receptor (EGFR) therapy is of interest as a mechanism for acquired resistance to anti-EGFR treatment. Plasma analysis of circulating tumor (ct) DNA is a minimally invasive and highly sensitive method to determine RAS mutational status. This biomarker analysis of the global phase III ASPECCT study used next-generation sequencing to detect expanded RAS ctDNA mutations in panitumumab-treated patients. Plasma samples collected at baseline and posttreatment were analyzed categorically for the presence of RAS mutations by the Plasma Select -R™ 64-gene panel at 0.1% sensitivity. Among panitumumab-treated patients with evaluable plasma samples at baseline (n = 238), 188 (79%) were wild-type (WT) RAS, and 50 (21%) were mutant RAS Of the 188 patients with baseline ctDNA WT RAS status, 164 had evaluable posttreatment results with a 32% rate of emergent RAS mutations. The median overall survival (OS) for WT and RAS mutant status by ctDNA at baseline was 13.7 (95% confidence interval: 11.5-15.4) and 7.9 months (6.4-9.6), respectively ( P < 0.0001). Clinical outcomes were not significantly different between patients with and without emergent ctDNA RAS mutations. Although patients with baseline ctDNA RAS mutations had worse outcomes than patients who were WT RAS before initiating treatment, emergent ctDNA RAS mutations were not associated with less favorable patient outcomes in panitumumab-treated patients. Further research is needed to determine a clinically relevant threshold for baseline and emergent ctDNA RAS mutations. Copyright ©2018, American Association for Cancer Research.

High Intra- and Inter-Tumoral Heterogeneity of RAS Mutations in Colorectal Cancer

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Marion Jeantet

2016-12-01

Full Text Available Approximately 30% of patients with wild type RAS metastatic colorectal cancer are non-responders to anti-epidermal growth factor receptor monoclonal antibodies (anti-EGFR mAbs, possibly due to undetected tumoral subclones harboring RAS mutations. The aim of this study was to analyze the distribution of RAS mutations in different areas of the primary tumor, metastatic lymph nodes and distant metastasis. A retrospective cohort of 18 patients with a colorectal cancer (CRC was included in the study. Multiregion analysis was performed in 60 spatially separated tumor areas according to the pathological tumor node metastasis (pTNM staging and KRAS, NRAS and BRAF mutations were tested using pyrosequencing. In primary tumors, intra-tumoral heterogeneity for RAS mutation was found in 33% of cases. Inter-tumoral heterogeneity for RAS mutation between primary tumors and metastatic lymph nodes or distant metastasis was found in 36% of cases. Moreover, 28% of tumors had multiple RAS mutated subclones in the same tumor. A high proportion of CRCs presented intra- and/or inter-tumoral heterogeneity, which has relevant clinical implications for anti-EGFR mAbs prescription. These results suggest the need for multiple RAS testing in different parts of the same tumor and/or more sensitive techniques.

Human hypervariable sequences in risk assessment: rare Ha-ras alleles in cancer patients

International Nuclear Information System (INIS)

Krontiris, T.G.; DiMartino, N.A.; Mitcheson, H.D.; Lonergan, J.A.; Begg, C.; Parkinson, D.R.

1987-01-01

A variable tandem repeat (VTR) is responsible for the hyperallelism one kilobase 3' to the human c-Ha-ras-1 (Ha-ras) gene. Thirty-two distinct restriction fragments, comprising 3 allelic classes by frequency of occurrence, have thus far been detected in a sample size of approximately 800 caucasians. Rare Ha-ras alleles, 21 in all, are almost exclusively confined to the genomes of cancer patients. From their data the authors have computed the relative cancer risk associated with possession of a rare Ha-ras allele to be 27. To understand the molecular basis for this phenomenon, they have begun to clone Ha-ras fragments from nontumor DNA of cancer patients. They report here the weak activation, as detected by transfection and transformation of NIH 3T3 mouse cells, of two Ha-ras genes which were obtained from lymphocyte DNA of a melanoma patient. They have mapped the regions that confer this transforming activity to the fragment containing the VTR in one Ha-ras clone and the fragment containing gene coding sequences in the other

The nitric oxide-sensitive p21Ras-ERK pathway mediates S-nitrosoglutathione-induced apoptosis

International Nuclear Information System (INIS)

Tsujita, Maristela; Batista, Wagner L.; Ogata, Fernando T.; Stern, Arnold; Monteiro, Hugo P.; Arai, Roberto J.

2008-01-01

p21Ras protein plays a critical role in cellular signaling that induces either cell cycle progression or apoptosis. Nitric oxide (NO) has been consistently reported to activate p21Ras through the redox sensitive cysteine residue (118). In this study, we demonstrated that the p21Ras-ERK pathway regulates THP-1 monocyte/macrophage apoptosis induced by S-nitrosoglutathione (SNOG). This was apparent from studies in THP-1 cells expressing NO-insensitive p21Ras (p21Ras C118S ) where the pro-apoptotic action of SNOG was almost abrogated. Three major MAP kinase pathways (ERK, JNK, and p38) that are downstream to p21Ras were investigated. It was observed that only the activation of ERK1/2 MAP kinases by SNOG in THP-1 cells was attributable to p21Ras. The inhibition of the ERK pathway by PD98059 markedly attenuated apoptosis in SNOG-treated THP-1 cells, but had a marginal effect on SNOG-treated THP-1 cells expressing NO-insensitive p21Ras. The inhibition of the JNK and p38 pathways by selective inhibitors had no marked effects on the percentage of apoptosis. The induction of p21Waf1 expression by SNOG was observed in THP-1 cells harboring mutant and wild-type p21Ras, however in cells expressing mutant Ras, the expression of p21Waf1 was significantly attenuated. The treatment of THP-1 cells expressing wild-type p21Ras with PD98059 resulted in significant attenuation of p21Waf1 expression. These results indicate that the redox sensitive p21Ras-ERK pathway plays a critical role in sensing and delivering the pro-apoptotic signaling mediated by SNOG

Generation of a double binary transgenic zebrafish model to study myeloid gene regulation in response to oncogene activation in melanocytes.

Science.gov (United States)

Kenyon, Amy; Gavriouchkina, Daria; Zorman, Jernej; Chong-Morrison, Vanessa; Napolitani, Giorgio; Cerundolo, Vincenzo; Sauka-Spengler, Tatjana

2018-04-06

A complex network of inflammatory genes is closely linked to somatic cell transformation and malignant disease. Immune cells and their associated molecules are responsible for detecting and eliminating cancer cells as they establish themselves as the precursors of a tumour. By the time a patient has a detectable solid tumour, cancer cells have escaped the initial immune response mechanisms. Here, we describe the development of a double binary zebrafish model that enables regulatory programming of the myeloid cells as they respond to oncogene-activated melanocytes to be explored, focussing on the initial phase when cells become the precursors of cancer. A hormone-inducible binary system allows for temporal control of expression of different Ras oncogenes ( NRas Q61K , HRas G12V and KRas G12V ) in melanocytes, leading to proliferation and changes in morphology of the melanocytes. This model was coupled to binary cell-specific biotagging models allowing in vivo biotinylation and subsequent isolation of macrophage or neutrophil nuclei for regulatory profiling of their active transcriptomes. Nuclear transcriptional profiling of neutrophils, performed as they respond to the earliest precursors of melanoma in vivo , revealed an intricate landscape of regulatory factors that may promote progression to melanoma, including Serpinb1l4, Fgf1, Fgf6, Cathepsin H, Galectin 1 and Galectin 3. The model presented here provides a powerful platform to study the myeloid response to the earliest precursors of melanoma. © 2018. Published by The Company of Biologists Ltd.

Literature review : performance of RAP/RAS mixes and new direction.

Science.gov (United States)

2014-04-01

In the last several years reclaimed asphalt pavement (RAP) and recycled asphalt shingles (RAS) have been : widely used in asphalt mixes in Texas. The use of RAP/RAS can significantly reduce the initial cost of : asphalt mixtures, conserve energy, and...

A Screen Identifies the Oncogenic Micro-RNA miR-378a-5p as a Negative Regulator of Oncogene-Induced Senescence

DEFF Research Database (Denmark)

Kooistra, Susanne Marije; Rudkjær, Lise Christine; Lees, Michael James

2014-01-01

Oncogene-induced senescence (OIS) can occur in response to hyperactive oncogenic signals and is believed to be a fail-safe mechanism protecting against tumorigenesis. To identify new factors involved in OIS, we performed a screen for microRNAs that can overcome or inhibit OIS in human diploid fib...

RAS in Pregnancy and Preeclampsia and Eclampsia

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M. Rodriguez

2012-01-01

Full Text Available Preeclampsia is a common disease of pregnancy characterized by the presence of hypertension and commitment of many organs, including the brain, secondary to generalized endothelial dysfunction. Its etiology is not known precisely, but it involved several factors, highlighting the renin angiotensin system (RAS, which would have an important role in the origin of multisystem involvement. This paper reviews the evidence supporting the involvement of RAS in triggering the disease, in addition to the components of this system that would be involved and how it eventually produces brain engagement.

BMP suppresses PTEN expression via RAS/ERK signaling.

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Beck, Stayce E; Carethers, John M

2007-08-01

Bone morphogenetic protein (BMP), a member of the transforming growth factor beta family, classically utilizes the SMAD signaling pathway for its growth suppressive effects,and loss of this signaling cascade may accelerate cell growth. In the colon cancer predisposition syndrome Juvenile Polyposis, as well as in the late progression stages of nonsyndromic colorectal cancers, SMAD4 function is typically abrogated. Here, we utilized the SMAD4-null SW480 colon cancer cell line to examine BMPs effect on a potential target gene, PTEN, and how its expression might be regulated. Initial treatment of the SMAD4-null cells with BMP resulted in mild growth suppression, but with prolonged exposure to BMP, the cells become growth stimulatory, which coincided with observed decreases in transcription and translation of PTEN, and with corresponding increases in phospho-AKT protein levels. BMP-induced PTEN suppression was mediated via the RAS/ERK pathway, as pharmacologic inhibition of RAS/ERK, or interference with protein function in the cytosol by DN-RAS prevented BMP-induced growth promotion and changes in PTEN levels, as did treatment with noggin, a BMP ligand inhibitor. Thus, BMP downregulates PTEN via RAS/ERK in a SMAD4-null environment that contributes to cell growth, and constitutes a SMAD4-independent but BMP-responsive signaling pathway.

Combined targeting of BRAF and CRAF or BRAF and PI3K effector pathways is required for efficacy in NRAS mutant tumors.

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Bijay S Jaiswal

Full Text Available BACKGROUND: Oncogenic RAS is a highly validated cancer target. Attempts at targeting RAS directly have so far not succeeded in the clinic. Understanding downstream RAS-effectors that mediate oncogenesis in a RAS mutant setting will help tailor treatments that use RAS-effector inhibitors either alone or in combination to target RAS-driven tumors. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we have investigated the sufficiency of targeting RAS-effectors, RAF, MEK and PI3-Kinase either alone or in combination in RAS mutant lines, using an inducible shRNA in vivo mouse model system. We find that in colon cancer cells harboring a KRAS(G13D mutant allele, knocking down KRAS alone or the RAFs in combination or the RAF effectors, MEK1 and MEK2, together is effective in delaying tumor growth in vivo. In melanoma cells harboring an NRAS(Q61L or NRAS(Q61K mutant allele, we find that targeting NRAS alone or both BRAF and CRAF in combination or both BRAF and PIK3CA together showed efficacy. CONCLUSION/SIGNIFICANCE: Our data indicates that targeting oncogenic NRAS-driven melanomas require decrease in both pERK and pAKT downstream of RAS-effectors for efficacy. This can be achieved by either targeting both BRAF and CRAF or BRAF and PIK3CA simultaneously in NRAS mutant tumor cells.

A family affair: A Ral-exocyst-centered network links Ras, Rac, Rho signaling to control cell migration.

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Zago, Giulia; Biondini, Marco; Camonis, Jacques; Parrini, Maria Carla

2017-05-12

Cell migration is central to many developmental, physiologic and pathological processes, including cancer progression. The Ral GTPases (RalA and RalB) which act down-stream the Ras oncogenes, are key players in the coordination between membrane trafficking and actin polymerization. A major direct effector of Ral, the exocyst complex, works in polarized exocytosis and is at the center of multiple protein-protein interactions that support cell migration by promoting protrusion formation, front-rear polarization, and extra-cellular matrix degradation. In this review we describe the recent advancements in deciphering the molecular mechanisms underlying this role of Ral via exocyst on cell migration. Among others, we will discuss the recently identified cross-talk between Ral and Rac1 pathways: exocyst binds to a negative regulator (the RacGAP SH3BP1) and to the major effector (the Wave Regulatory Complex, WRC) of Rac1, the master regulator of protrusions. Next challenge will be to better characterize the dynamics in space and in time of these molecular interplays, to better understand the pleiotropic functions of Ral in both normal and cancer cells.

Transcriptional Profile of Ki-Ras-Induced Transformation of Thyroid Cells

DEFF Research Database (Denmark)

Visconti, Roberta; Federico, Antonella; Coppola, Valeria

2007-01-01

Abstract In the last years, an increasing number of experiments has provided compelling evidence for a casual role of Ras protein mutations, resulting in their constitutive activation, in thyroid carcinogenesis. However, despite the clear involvement of Ras proteins in thyroid carcinogenesis, the...

Development of a High-Throughput Gene Expression Screen for Modulators of RAS-MAPK Signaling in a Mutant RAS Cellular Context.

Science.gov (United States)

Severyn, Bryan; Nguyen, Thi; Altman, Michael D; Li, Lixia; Nagashima, Kumiko; Naumov, George N; Sathyanarayanan, Sriram; Cook, Erica; Morris, Erick; Ferrer, Marc; Arthur, Bill; Benita, Yair; Watters, Jim; Loboda, Andrey; Hermes, Jeff; Gilliland, D Gary; Cleary, Michelle A; Carroll, Pamela M; Strack, Peter; Tudor, Matt; Andersen, Jannik N

2016-10-01

The RAS-MAPK pathway controls many cellular programs, including cell proliferation, differentiation, and apoptosis. In colorectal cancers, recurrent mutations in this pathway often lead to increased cell signaling that may contribute to the development of neoplasms, thereby making this pathway attractive for therapeutic intervention. To this end, we developed a 26-member gene signature of RAS-MAPK pathway activity utilizing the Affymetrix QuantiGene Plex 2.0 reagent system and performed both primary and confirmatory gene expression-based high-throughput screens (GE-HTSs) using KRAS mutant colon cancer cells (SW837) and leveraging a highly annotated chemical library. The screen achieved a hit rate of 1.4% and was able to enrich for hit compounds that target RAS-MAPK pathway members such as MEK and EGFR. Sensitivity and selectivity performance measurements were 0.84 and 1.00, respectively, indicating high true-positive and true-negative rates. Active compounds from the primary screen were confirmed in a dose-response GE-HTS assay, a GE-HTS assay using 14 additional cancer cell lines, and an in vitro colony formation assay. Altogether, our data suggest that this GE-HTS assay will be useful for larger unbiased chemical screens to identify novel compounds and mechanisms that may modulate the RAS-MAPK pathway. © 2016 Society for Laboratory Automation and Screening.

The Human Cytomegalovirus Strain DB Activates Oncogenic Pathways in Mammary Epithelial Cells

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Amit Kumar

2018-04-01

Full Text Available Background: Human cytomegalovirus (HCMV establishes a persistent life-long infection and increasing evidence indicates HCMV infection can modulate signaling pathways associated with oncogenesis. Breast milk is an important route of HCMV transmission in humans and we hypothesized that mammary epithelial cells could be one of the main cellular targets of HCMV infection. Methods: The infectivity of primary human mammary epithelial cells (HMECs was assessed following infection with the HCMV-DB strain, a clinical isolate with a marked macrophage-tropism. The impact of HCMV-DB infection on expression of p53 and retinoblastoma proteins, telomerase activity and oncogenic pathways (c-Myc, Akt, Ras, STAT3 was studied. Finally the transformation of HCMV-DB infected HMECs was evaluated using soft agar assay. CTH cells (CMV Transformed HMECs were detected in prolonged cultures of infected HMECs. Tumor formation was observed in NOD/SCID Gamma (NSG mice injected with CTH cells. Detection of long non coding RNA4.9 (lncRNA4.9 gene was assessed in CTH cells, tumors isolated from xenografted NSG mice and biopsies of patients with breast cancer using qualitative and quantitative PCR. Results: We found that HCMV, especially a clinical strain named HCMV-DB, infects HMECs in vitro. The clinical strain HCMV-DB replicates productively in HMECs as evidenced by detection of early and late viral transcripts and proteins. Following infection of HMECs with HCMV-DB, we observed the inactivation of retinoblastoma and p53 proteins, the activation of telomerase activity, the activation of the proto-oncogenes c-Myc and Ras, the activation of Akt and STAT3, and the upregulation of cyclin D1 and Ki67 antigen. Colony formation was observed in soft agar seeded with HCMV-DB-infected HMECs. Prolonged culture of infected HMECs resulted in the development of clusters of spheroid cells that we called CTH cells (CMV Transformed HMECs. CTH cells when injected in NOD/SCID Gamma (NSG mice

DNA damage and repair in oncogenic transformation by heavy ion radiation

Science.gov (United States)

Yang, T. C.; Mei, M.; George, K. A.; Craise, L. M.

1996-01-01

Energetic heavy ions are present in galactic cosmic rays and solar particle events. One of the most important late effects in risk assessment is carcinogenesis. We have studied the carcinogenic effects of heavy ions at the cellular and molecular levels and have obtained quantitative data on dose-response curves and on the repair of oncogenic lesions for heavy particles with various charges and energies. Studies with repair inhibitors and restriction endonucleases indicated that for oncogenic transformation DNA is the primary target. Results from heavy ion experiments showed that the cross section increased with LET and reached a maximum value of about 0.02 micrometer2 at about 500 keV/micrometer. This limited size of cross section suggests that only a fraction of cellular genomic DNA is important in radiogenic transformation. Free radical scavengers, such as DMSO, do not give any effect on induction of oncogenic transformation by 600 MeV/u iron particles, suggesting most oncogenic damage induced by high-LET heavy ions is through direct action. Repair studies with stationary phase cells showed that the amount of reparable oncogenic lesions decreased with an increase of LET and that heavy ions with LET greater than 200 keV/micrometer produced only irreparable oncogenic damage. An enhancement effect for oncogenic transformation was observed in cells irradiated by low-dose-rate argon ions (400 MeV/u; 120 keV/micrometer). Chromosomal aberrations, such as translocation and deletion, but not sister chromatid exchange, are essential for heavy-ion-induced oncogenic transformation. The basic mechanism(s) of misrepair of DNA damage, which form oncogenic lesions, is unknown.

Machine Learning Detects Pan-cancer Ras Pathway Activation in The Cancer Genome Atlas

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Gregory P. Way

2018-04-01

Full Text Available Summary: Precision oncology uses genomic evidence to match patients with treatment but often fails to identify all patients who may respond. The transcriptome of these “hidden responders” may reveal responsive molecular states. We describe and evaluate a machine-learning approach to classify aberrant pathway activity in tumors, which may aid in hidden responder identification. The algorithm integrates RNA-seq, copy number, and mutations from 33 different cancer types across The Cancer Genome Atlas (TCGA PanCanAtlas project to predict aberrant molecular states in tumors. Applied to the Ras pathway, the method detects Ras activation across cancer types and identifies phenocopying variants. The model, trained on human tumors, can predict response to MEK inhibitors in wild-type Ras cell lines. We also present data that suggest that multiple hits in the Ras pathway confer increased Ras activity. The transcriptome is underused in precision oncology and, combined with machine learning, can aid in the identification of hidden responders. : Way et al. develop a machine-learning approach using PanCanAtlas data to detect Ras activation in cancer. Integrating mutation, copy number, and expression data, the authors show that their method detects Ras-activating variants in tumors and sensitivity to MEK inhibitors in cell lines. Keywords: Gene expression, machine learning, Ras, NF1, KRAS, NRAS, HRAS, pan-cancer, TCGA, drug sensitivity

Active Erk Regulates Microtubule Stability in H-ras-Transformed Cells

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Rene E. Harrison

2001-01-01

Full Text Available Increasing evidence suggests that activated erk regulates cell functions, at least in part, by mechanisms that do not require gene transcription. Here we show that the map kinase, erk, decorates microtubules (MTs and mitotic spindles in both parental and mutant active rastransfected 10T1 /2 fibroblasts and MCF10A breast epithelial cells. Approximately 20% of total cellular erk decorated MTs in both cell lines. A greater proportion of activated erk was associated with MTs in the presence of mutant active H-ras than in parental cells. Activation of erk by the ras pathway coincided with a decrease in the stability of MT, as detected by a stability marker. The MKK1 inhibitor, PD98059 and transfection of a dominant negative MKK1 blocked ras-induced instability of MTs but did not modify the association of erk with MTs or affect MT stability of the parental cells. These results indicate that the subset of active erk kinase that associates with MTs contributes to their instability in the presence of a mutant active ras. The MT-associated subset of active erk likely contributes to the enhanced invasive and proliferative abilities of cells containing mutant active H-ras.

Tumorigenesis of K-ras mutation in human endometrial carcinoma via upregulation of estrogen receptor.

Science.gov (United States)

Tu, Zheng; Gui, Liming; Wang, Jianliu; Li, Xiaoping; Sun, Pengming; Wei, Lihui

2006-05-01

To investigate the tumorigenesis of mutant [12Asp]-K-ras in endometrial carcinoma and its relationship with ER. We constructed pcDI-[12Asp]K-ras4B by inserting full-length [12Asp]K-ras4B from human endometrial carcinoma Hec-1A cells, into pcDI vector. Cell proliferation of NIH3T3 after transfection with pcDI-[12Asp]K-ras4B was measured by MTT assay. The cell transformation was determined by colony formation and tumor nodule development. [12Asp]-K-ras4B-NIH3T3 cells were transfected with constitutively active pCMV-RafCAAX and dominant-negative pCMV-RafS621A. Cell growth was measured by MTT assay and [3H]thymidine incorporation. After transfected with pcDI-[12Asp]K-ras4B or pCMV-RafS621A, the cells were harvested for Western blot and reporter assay to determine the expression and transcriptional activity of ERalpha and ERbeta, respectively. [12Asp]-K-ras4B enhanced NIH3T3 cells proliferation after 48 h post-transfection (P ras4B-NIH3T3 cells (13.48%) than pcDI-NIH3T3 (4.26%) or untreated NIH3T3 (2.33%). The pcDI-[12Asp]-K-ras4B-NIH3T3 cells injected to the nude mice Balb/C developed tumor nodules with poor-differentiated cells after 12 days. An increase of ERalpha and ERbeta was observed in pcDI-[12Asp]-K-ras4B-NIH3T3 cells. RafS621A downregulated ERalpha and ERbeta expression. Estrogen induced the ER transcriptional activity by 5-fold in pcDI-NIH3T3 cells, 13-fold in pcDI-[12Asp]K-ras4B-NIH3T3 and 19-fold in HEC-1A. RafS621A suppressed the ER transcriptional activity. K-ras mutation induces tumorigenesis in endometrium, and this malignant transformation involves Raf signaling pathway and ER.

Analisis Kelembagaan Rantai Pasok Telur Ayam Ras Peternakan Rakyat Di Jawa Barat

OpenAIRE

Sejati, Wahyuning K

2011-01-01

Dalam konteks pengembangan pasar komoditas dan peningkatan kesejahteraan peternak rakyat perlu dipahami secara baik karakteristik dan kelembagaan petani, pemasok, dan pasar. Tulisan ini bertujuan untuk menganalisis kelembagaan rantai pasok peternak-pemasok-pasar komoditas telur ayam ras yang meliputi: (1) identifikasi kelembagaan peternakan rakyat ayam ras petelur; (2) analisis kelembagaan rantai pasok komoditas telur; dan (3) antisipasi kelembagaan introduksi rantai pasok telur ayam ras pete...

Identification of H-Ras-Specific Motif for the Activation of Invasive Signaling Program in Human Breast Epithelial Cells

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Hae-Young Yong

2011-02-01

Full Text Available Increased expression and/or activation of H-Ras are often associated with tumor aggressiveness in breast cancer. Previously, we showed that H-Ras, but not N-Ras, induces MCF10A human breast epithelial cell invasion and migration, whereas both H-Ras and N-Ras induce cell proliferation and phenotypic transformation. In an attempt to determine the sequence requirement directing the divergent phenotype induced by H-Ras and N-Ras with a focus on the induction of human breast cell invasion, we investigated the structural and functional relationships between H-Ras and N-Ras using domain-swap and site-directed mutagenesis approaches. Here, we report that the hypervariable region (HVR, consisting of amino acids 166 to 189 in H-Ras, determines the invasive/migratory signaling program as shown by the exchange of invasive phenotype by swapping HVR sequences between H-Ras and N-Ras. We also demonstrate that the H-Ras-specific additional palmitoylation site at Cys184 is not responsible for the signaling events that distinguish between H-Ras and N-Ras. Importantly, this work identifies the C-terminal HVR, especially the flexible linker domain with two consecutive proline residues Pro173 and Pro174, as a critical domain that contributes to activation of H-Ras and its invasive potential in human breast epithelial cells. The present study sheds light on the structural basis for the Ras isoform-specific invasive program of breast epithelial cells, providing information for the development of agents that specifically target invasion-related H-Ras pathways in human cancer.

A mouse model for Costello syndrome reveals an Ang II–mediated hypertensive condition

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Schuhmacher, Alberto J.; Guerra, Carmen; Sauzeau, Vincent; Cañamero, Marta; Bustelo, Xosé R.; Barbacid, Mariano

2008-01-01

Germline activation of H-RAS oncogenes is the primary cause of Costello syndrome (CS), a neuro-cardio-facio-cutaneous developmental syndrome. Here we describe the generation of a mouse model of CS by introduction of an oncogenic Gly12Val mutation in the mouse H-Ras locus using homologous recombination in ES cells. Germline expression of the endogenous H-RasG12V oncogene, even in homozygosis, resulted in hyperplasia of the mammary gland. However, development of tumors in these mice was rare. H-RasG12V mutant mice closely phenocopied some of the abnormalities observed in patients with CS, including facial dysmorphia and cardiomyopathies. These mice also displayed alterations in the homeostasis of the cardiovascular system, including development of systemic hypertension, extensive vascular remodeling, and fibrosis in both the heart and the kidneys. This phenotype was age dependent and was a consequence of the abnormal upregulation of the renin–Ang II system. Treatment with captopril, an inhibitor of Ang II biosynthesis, prevented development of the hypertension condition, vascular remodeling, and heart and kidney fibrosis. In addition, it partially alleviated the observed cardiomyopathies. These mice should help in elucidating the etiology of CS symptoms, identifying additional defects, and evaluating potential therapeutic strategies. PMID:18483625

H-ras oncogene-transformed human bronchial epithelial cells (TBE-1) secrete a single metalloprotease capable of degrading basement membrane collagen

International Nuclear Information System (INIS)

Collier, I.E.; Wilhelm, S.M.; Eisen, A.Z.

1988-01-01

H-ras transformed human bronchial epithelial cells (TBE-1) secrete a single major extracellular matrix metalloprotease which is not found in the normal parental cells. The enzyme is secreted in a latent form which can be activated to catalyze the cleavage of the basement membrane macromolecule type IV collagen. The substrates in their order of preference are: gelatin, type IV collagen, type V collagen, fibronectin, and type VII collagen; but the enzyme does not cleave the interstitial collagens or laminin. This protease is identical to gelatinase isolated from normal human skin explants, normal human skin fibroblasts, and SV40-transformed human lung fibroblasts. Based on this ability to initiate the degradation of type IV collagen in a pepsin-resistant portion of the molecule, it will be referred to as type IV collagenase. This enzyme is most likely the human analog of type IV collagenase detected in several rodent tumors. Type IV collagenase consists of three domains. Type IV collagenase represents the third member of a newly recognized gene family coding for secreted extracellular matrix metalloproteases, which includes interstitial fibroblast collagenase and stromelysin

Subcellular Distribution of S-Nitrosylated H-Ras in Differentiated and Undifferentiated PC12 Cells during Hypoxia.

Science.gov (United States)

Barbakadze, Tamar; Goloshvili, Galina; Narmania, Nana; Zhuravliova, Elene; Mikeladze, David

2017-10-01

Hypoxia or exposure to excessive reactive oxygen or nitrogen species could induce S-nitrosylation of various target proteins, including GTPases of the Ras-superfamily. Under hypoxic conditions, the Ras-protein is translocated to the cytosol and interacts with the Golgi complex, endoplasmic reticulum, mitochondria. The mobility/translocation of Ras depend on the cells oxidative status. However, the importance of relocated Snitrosylated- H-Ras (NO-H-Ras) in proliferation/differentiation processes is not completely understood. We have determined the content of soluble- and membrane-bound-NO-HRas in differentiated (D) and undifferentiated (ND) rat pheochromocytoma (PC12) cells under hypoxic and normoxic conditions. In our experimental study, we analyzed NO-H-Ras levels under hypoxic/normoxic conditions in membrane and soluble fractions of ND and D PC12 cells with/without nitric oxide donor, sodium nitroprusside (SNP) treatment. Cells were analyzed by the S-nitrosylated kit, immunoprecipitation, and Western blot. We assessed the action of NO-H-Ras on oxidative metabolism of isolated mitochondria by determining mitochondrial hydrogen peroxide generation via the scopoletin oxidation method and ATPproduction as estimated by the luminometric method. Hypoxia did not influence nitrosylation of soluble H-Ras in ND PC12 cells. Under hypoxic conditions, the nitrosylation of soluble-H-Ras greatly decreased in D PC12 cells. SNP didn't change the levels of nitrosylation of soluble-H-Ras, in either hypoxic or normoxic conditions. On the other hand, hypoxia, per se, did not affect the nitrosylation of membrane-bound-H-Ras in D and ND PC12 cells. SNP-dependent nitrosylation of membrane-bound-H-Ras greatly increased in D PC12 cells. Both unmodified normal and mutated H-Ras enhanced the mitochondrial synthesis of ATP, whereas the stimulatory effects on ATP synthesis were eliminated after S-nitrosylation of H-Ras. According to the results, it may be proposed that hypoxia can decrease S

Experiment list: SRX017560 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available s=Normal 10468397,71.3,11.7,1639 GSM508794: BJ p53KD +RAS IPp53 source_name=BJ primary foreskin fibroblasts, p53 knock...down IP || cell line=BJ || cell type=foreskin fibroblasts || oncogene expression=RasV12 || vector=p53 knock

Experiment list: SRX017558 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available s=Normal 10966699,74.9,8.3,1410 GSM508792: BJ BRD7KD +RAS IPBRD7 source_name=BJ primary foreskin fibroblasts, BRD7 knock...down IP || cell line=BJ || cell type=foreskin fibroblasts || oncogene expression=RasV12 || vector=BRD7 knock

[Expressions of Ras and Sos1 in epithelial ovarian cancer tissues and their clinical significance].

Science.gov (United States)

Xiao, Zheng-Hua; Linghu, Hua; Liu, Qian-Fen

2016-11-20

To detect the expressions of Ras and Sos1 proteins in human epithelial ovarian cancer (EOC) tissues and explore their correlation with the clinicopathological features of the patients. The expressions of Ras and Sos1 proteins were detected immunohistochemically in 62 EOC tissues, 5 borderline ovarian cancer tissues, 15 benign epithelial ovarian neoplasm tissues, and 18 normal ovarian tissues. The EOC tissues showed significantly higher expression levels of both Ras and Sos1 than the other tissues tested (Ptissues, Ras and Sos1 proteins were expressed mostly on the cell membrane and in the cytoplasm. The expression level of Ras was correlated with pathological types of the tumor (Ptissue-specific variation of Ras expression can lend support to a specific diagnosis of ovarian serous adenocarcinoma. The association of Ras and Sos1 protein expression with the tumor-free survival time of the patients awaits further investigation with a larger sample size.

Aberrant status and clinicopathologic characteristic associations of 11 target genes in 1,321 Chinese patients with lung adenocarcinoma.

Science.gov (United States)

Zhao, Mengnan; Zhan, Cheng; Li, Ming; Yang, Xiaodong; Yang, Xinyu; Zhang, Yong; Lin, Miao; Xia, Yifeng; Feng, Mingxiang; Wang, Qun

2018-01-01

The aberrant status of target genes and their associations with clinicopathologic characteristics are still unclear in primary lung adenocarcinoma. The common mutations and translocations of nine target genes were evaluated in 1,247 specimens of surgically-resected primary lung adenocarcinoma. Immunohistochemistry was used to analyze the expressions of programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1) in 731 specimens. The frequency of the aberrations and their associations with clinicopathologic characteristics were analyzed. Overall, 952 (76.3%) of 1,247 patients harbored at least one target mutation or translocation: epidermal growth factor receptor ( EGFR ) (729, 58.5%), v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog ( KRAS ) (83, 6.7%), human epidermal growth factor receptor 2 ( HER2 ) (82, 6.6%), anaplastic lymphoma kinase ( ALK) (23, 1.8%), phosphoinositide-3-kinase catalytic alpha polypeptide ( PIK3CA ) (20, 1.6%), Ret proto-oncogene RET (15, 1.2%), ROS proto-oncogene 1 receptor tyrosine kinase ( ROS1 ) (12, 1.0%), B-raf proto-oncogene ( BRAF ) (9, 0.7%), neuroblastoma RAS viral (v-ras) oncogene homolog ( NRAS ) (3, 0.2%). Fourteen (1.9%) of 731 patients were PD-1 positive and 95 (13.0%) were PD-L1 positive in tumor cells. In men and smokers, there were more frequent KRAS mutations (both Ppatients, while HER2 (Ppatients with EGFR mutations (all Ppatients with primary lung adenocarcinoma harbored target gene aberrations. The frequency of each alteration differed in patients depending on clinicopathologic characteristics.

K-ras2 Activation and Genome Instability Increase Proliferation and Size of FAP Adenomas

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Anna Rapallo

1999-01-01

Full Text Available The possible role of K‐ras2 mutations and aneuploidy toward increase of proliferation and adenoma size in Familial Adenomatous Polyposis (FAP adenomas is not known. The present study addresses these issues by investigating 147 colorectal adenomas obtained from four FAP patients. The majority of adenomas had size lower than or equal to 10 mm (86%, low grade dysplasia (63%, and were preferentially located in the right colon (60%. Normal mucosa samples were obtained from 19 healthy donors. Three synchronous adenocarcinomas were also investigated. K‐ras2 mutation spectrum was analysed by PCR and Sequence Specific Oligonucleotide (SSO hybridization, while flow cytometry (FCM was used for evaluating degree of DNA ploidy and S‐phase fraction. Overall, incidences of K‐ras2 mutations, DNA aneuploidy and high S‐phase values (>7.2% were 6.6%, 5.4% and 10.5%, respectively. In particular, among the adenomas with size lower than 5 mm, K‐ras2 mutation and DNA aneuploidy frequencies were only slightly above 1%. Statistically significant correlations were found between K‐ras2 and size, DNA ploidy and size and K‐ras2 and S‐phase (p. In particular, among the wild type K‐ras2 adenomas, high S‐phase values were detected in 8% of the cases versus 57% among the K‐ras2 mutated adenomas (p=0.0005. The present series of FAP adenomas indicates that K‐ras2 activation and gross genomic changes play a role toward a proliferative gain and tumour growth in size.

Bacillus subtilis Intramembrane Protease RasP Activity in Escherichia coli and In Vitro.

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Parrell, Daniel; Zhang, Yang; Olenic, Sandra; Kroos, Lee

2017-10-01

RasP is a predicted intramembrane metalloprotease of Bacillus subtilis that has been proposed to cleave the stress response anti-sigma factors RsiW and RsiV, the cell division protein FtsL, and remnant signal peptides within their transmembrane segments. To provide evidence for direct effects of RasP on putative substrates, we developed a heterologous coexpression system. Since expression of catalytically inactive RasP E21A inhibited expression of other membrane proteins in Escherichia coli , we added extra transmembrane segments to RasP E21A, which allowed accumulation of most other membrane proteins. A corresponding active version of RasP appeared to promiscuously cleave coexpressed membrane proteins, except those with a large periplasmic domain. However, stable cleavage products were not observed, even in clpP mutant E. coli Fusions of transmembrane segment-containing parts of FtsL and RsiW to E. coli maltose-binding protein (MBP) also resulted in proteins that appeared to be RasP substrates upon coexpression in E. coli , including FtsL with a full-length C-terminal domain (suggesting that prior cleavage by a site 1 protease is unnecessary) and RsiW designed to mimic the PrsW site 1 cleavage product (suggesting that further trimming by extracytoplasmic protease is unnecessary). Purified RasP cleaved His 6 -MBP-RsiW(73-118) in vitro within the RsiW transmembrane segment based on mass spectrometry analysis, demonstrating that RasP is an intramembrane protease. Surprisingly, purified RasP failed to cleave His 6 -MBP-FtsL(23-117). We propose that the lack of α-helix-breaking residues in the FtsL transmembrane segment creates a requirement for the membrane environment and/or an additional protein(s) in order for RasP to cleave FtsL. IMPORTANCE Intramembrane proteases govern important signaling pathways in nearly all organisms. In bacteria, they function in stress responses, cell division, pathogenesis, and other processes. Their membrane-associated substrates are

A role for HB-EGF/EGFR and MAPK signaling in the development of intestinal serrated polyps in mice

NARCIS (Netherlands)

Bongers, G.M.; Muniz, L. R.; Pacer, M.; Thirunarayanan, N.; Smit, M.J.; Reddy, E.P.; Mayer, L.; Furtado, G.C.; Harpaz, N.; Lira, SA

2012-01-01

BACKGROUND & AIMS: Epithelial cancers can be initiated by activating mutations in components of the mitogen-activated protein kinase signaling pathway such as v-raf murine sarcoma viral oncogene homolog B1 (BRAF), v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS), or epidermal growth

Epac activation sensitizes rat sensory neurons via activation of Ras

Science.gov (United States)

Shariati, Behzad; Thompson, Eric L.; Nicol, Grant D.; Vasko, Michael R.

2015-01-01

Guanine nucleotide exchange factors directly activated by cAMP (Epacs) have emerged as important signaling molecules mediating persistent hypersensitivity in animal models of inflammation, by augmenting the excitability of sensory neurons. Although Epacs activate numerous downstream signaling cascades, the intracellular signaling which mediates Epac-induced sensitization of capsaicin-sensitive sensory neurons remains unknown. Here, we demonstrate that selective activation of Epacs with 8-CPT-2′-O-Me-cAMP-AM (8CPT-AM) increases the number of action potentials (APs) generated by a ramp of depolarizing current and augments the evoked release of calcitonin gene-related peptide (CGRP) from isolated rat sensory neurons. Internal perfusion of capsaicin-sensitive sensory neurons with GDP-βS, substituted for GTP, blocks the ability of 8CPT-AM to increase AP firing, demonstrating that Epac-induced sensitization is G-protein dependent. Treatment with 8CPT-AM activates the small G-proteins Rap1 and Ras in cultures of sensory neurons. Inhibition of Rap1, by internal perfusion of a Rap1-neutralizing antibody or through a reduction in the expression of the protein using shRNA does not alter the Epac-induced enhancement of AP generation or CGRP release, despite the fact that in most other cell types, Epacs act as Rap-GEFs. In contrast, inhibition of Ras through expression of a dominant negative Ras (DN-Ras) or through internal perfusion of a Ras-neutralizing antibody blocks the increase in AP firing and attenuates the increase in the evoked release of CGRP induced by Epac activation. Thus, in this subpopulation of nociceptive sensory neurons, it is the novel interplay between Epacs and Ras, rather than the canonical Epacs and Rap1 pathway, that is critical for mediating Epac-induced sensitization. PMID:26596174

Epac activation sensitizes rat sensory neurons through activation of Ras.

Science.gov (United States)

Shariati, Behzad; Thompson, Eric L; Nicol, Grant D; Vasko, Michael R

2016-01-01

Guanine nucleotide exchange factors directly activated by cAMP (Epacs) have emerged as important signaling molecules mediating persistent hypersensitivity in animal models of inflammation, by augmenting the excitability of sensory neurons. Although Epacs activate numerous downstream signaling cascades, the intracellular signaling which mediates Epac-induced sensitization of capsaicin-sensitive sensory neurons remains unknown. Here, we demonstrate that selective activation of Epacs with 8-CPT-2'-O-Me-cAMP-AM (8CPT-AM) increases the number of action potentials (APs) generated by a ramp of depolarizing current and augments the evoked release of calcitonin gene-related peptide (CGRP) from isolated rat sensory neurons. Internal perfusion of capsaicin-sensitive sensory neurons with GDP-βS, substituted for GTP, blocks the ability of 8CPT-AM to increase AP firing, demonstrating that Epac-induced sensitization is G-protein dependent. Treatment with 8CPT-AM activates the small G-proteins Rap1 and Ras in cultures of sensory neurons. Inhibition of Rap1, by internal perfusion of a Rap1-neutralizing antibody or through a reduction in the expression of the protein using shRNA does not alter the Epac-induced enhancement of AP generation or CGRP release, despite the fact that in most other cell types, Epacs act as Rap-GEFs. In contrast, inhibition of Ras through expression of a dominant negative Ras (DN-Ras) or through internal perfusion of a Ras-neutralizing antibody blocks the increase in AP firing and attenuates the increase in the evoked release of CGRP induced by Epac activation. Thus, in this subpopulation of nociceptive sensory neurons, it is the novel interplay between Epacs and Ras, rather than the canonical Epacs and Rap1 pathway, that is critical for mediating Epac-induced sensitization. Copyright © 2015 Elsevier Inc. All rights reserved.

MRAS: A Close but Understudied Member of the RAS Family.

Science.gov (United States)

Young, Lucy C; Rodriguez-Viciana, Pablo

2018-01-08

MRAS is the closest relative to the classical RAS oncoproteins and shares most regulatory and effector interactions. However, it also has unique functions, including its ability to function as a phosphatase regulatory subunit when in complex with SHOC2 and protein phosphatase 1 (PP1). This phosphatase complex regulates a crucial step in the activation cycle of RAF kinases and provides a key coordinate input required for efficient ERK pathway activation and transformation by RAS. MRAS mutations rarely occur in cancer but deregulated expression may play a role in tumorigenesis in some settings. Activating mutations in MRAS (as well as SHOC2 and PP1) do occur in the RASopathy Noonan syndrome, underscoring a key role for MRAS within the RAS-ERK pathway. MRAS also has unique roles in cell migration and differentiation and has properties consistent with a key role in the regulation of cell polarity. Further investigations should shed light on what remains a relatively understudied RAS family member. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

Watershed Analysis with the Hydrologic Engineering Center's River Analysis System (HEC-RAS)

National Research Council Canada - National Science Library

Goodell, Christopher R; Brunner, Gary W

2004-01-01

The objectives of this document are to provide a general description of the HEC-RAS model, its capabilities and limitations, data requirements, traditional and innovative methods for HEC-RAS hydraulic...

Genetic modelling of PIM proteins in cancer: proviral tagging, cooperation with oncogenes, tumor suppressor genes and carcinogens.

Directory of Open Access Journals (Sweden)

Enara eAguirre

2014-05-01

Full Text Available The PIM proteins, which were initially discovered as proviral insertion sites in Moloney murine leukemia virus infection, are a family of highly homologous serine/threonine kinases that have been reported to be overexpressed in hematological malignancies and solid tumors. The PIM proteins have also been associated with metastasis and overall treatment responses and implicated in the regulation of apoptosis, metabolism, the cell cycle, and homing and migration, which makes these proteins interesting targets for anticancer drug discovery. The use of retroviral insertional mutagenesis and refined approaches such as complementation tagging has allowed the identification of myc, pim and a third group of genes (including bmi1 and gfi1 as complementing genes in lymphomagenesis. Moreover, mouse modeling of human cancer has provided an understanding of the molecular pathways that are involved in tumor initiation and progression at the physiological level. In particular, genetically modified mice have allowed researchers to further elucidate the role of each of the Pim isoforms in various tumor types. PIM kinases have been identified as weak oncogenes because experimental overexpression in lymphoid tissue, prostate and liver induces tumors at a relatively low incidence and with a long latency. However, very strong synergistic tumorigenicity between Pim1/2 and c-Myc and other oncogenes has been observed in lymphoid tissues. Mouse models have also been used to study whether the inhibition of specific PIM isoforms is required to prevent carcinogen-induced sarcomas, indicating that the absence of Pim2 and Pim3 greatly reduces sarcoma growth and bone invasion; the extent of this effect is similar to that observed in the absence of all 3 isoforms. This review will summarize some of the animal models that have been used to understand the isoform-specific contribution of PIM kinases to tumorigenesis.

Ras promotes cell survival by antagonizing both JNK and Hid signals in the Drosophila eye.

Science.gov (United States)

Wu, Yue; Zhuang, Yuan; Han, Min; Xu, Tian; Deng, Kejing

2009-10-20

Programmed cell death, or apoptosis, is a fundamental physiological process during normal development or in pathological conditions. The activation of apoptosis can be elicited by numerous signalling pathways. Ras is known to mediate anti-apoptotic signals by inhibiting Hid activity in the Drosophila eye. Here we report the isolation of a new loss-of-function ras allele, rasKP, which causes excessive apoptosis in the Drosophila eye. This new function is likely to be mediated through the JNK pathway since the inhibition of JNK signalling can significantly suppress rasKP-induced apoptosis, whereas the removal of hid only weakly suppresses the phenotype. Furthermore, the reduction of JNK signalling together with the expression of the baculovirus caspase inhibitor p35, which blocks Hid activity, strongly suppresses the rasKP cell death. In addition, we find a strong correlation between rasKP-induced apoptosis in the eye disc and the activation of JNK signalling. In the Drosophila eye, Ras may protect cells from apoptosis by inhibiting both JNK and Hid activities. Surprisingly, reducing Ras activity in the wing, however, does not cause apoptosis but rather affects cell and organ size. Thus, in addition to its requirement for cell viability, Ras appears to mediate different biological roles depending on the developmental context and on the level of its expression.

Regional Sediment Management (RSM) Modeling Tools: Integration of Advanced Sediment Transport Tools into HEC-RAS

Science.gov (United States)

2014-06-01

sediment transport within the USACE HEC River Analysis System ( HEC - RAS ) software package and to determine its applicability to Regional Sediment...Management (RSM) challenges. HEC - RAS SEDIMENT MODELING BACKGROUND: HEC - RAS performs (1) one- dimensional (1D) steady and unsteady hydraulic river ...Albuquerque (SPA)), and recently, the USACE RSM Program. HEC - RAS is one of several hydraulic modeling codes available for river analysis in the

Oncogenic osteomalacia associated with soft tissue chondromyxoid fibroma

International Nuclear Information System (INIS)

Park, Jeong Mi; Woo, Young Kyun; Kang, Moo Il; Kang, Chang Suk; Hahn, Seong Tae

2001-01-01

Oncogenic osteomalacia is a rarely described clinical entity characterized by hypophosphatemia, phosphaturia, and a low concentration of 1,25-dihydroxyvitamin D 3 . It is most often associated with benign mesenchymal tumor and can be cured with surgical removal of the tumor. In this paper, we present a case of oncogenic osteomalacia caused by chondromyxoid fibroma in the soft tissue of the sole of the foot in a 56-year-old woman

Loss of RASSF1A Expression in Colorectal Cancer and Its Association with K-ras Status

Directory of Open Access Journals (Sweden)

Dan Cao

2013-01-01

Full Text Available Background. The RAS-association domain family 1 A (RASSF1A is a classical member of RAS effectors regulating cell proliferation and apoptosis. Loss of RASSF1A expression may shift the balance towards a growth-promoting effect without the necessity of activating K-ras mutations. Its potential association with K-ras mutations in colorectal cancer (CRC is unclear. Methods. RASSF1A expression was examined in normal mucosa, adenoma, and tumor tissues of colon and rectum, respectively. We examined the association of RASSF1A expression, mutations of K-ras, and EGFR status in 76 primary CRCs. The relationship between clinicopathological characteristics and RASSF1A expression was also analyzed. Results. RASSF1A expression level decreased progressively in normal mucosa, adenoma and, tumor tissues, and the loss of RASSF1A expression occurred more frequently in tumor tissues. Of 76 primary CRCs, loss of RASSF1A expression and/or K-ras mutations were detected in 77% cases. Loss of RASSF1A expression was more frequent in K-ras wild-type than in mutation cases (63% versus 32%, . Conclusions. Our study indicates that loss of RASSF1A may be involved in pathogenesis of CRC, its expression was found predominantly in K-ras wild-type CRCs, suggesting that it may be another way of affecting RAS signaling, in addition to K-ras mutations.

EGFR immunoexpression, RAS immunoexpression and their effects on survival in lung adenocarcinoma cases.

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Gundogdu, Ahmet Gokhan; Onder, Sevgen; Firat, Pinar; Dogan, Riza

2014-06-01

The impacts of epidermal growth factor receptor (EGFR) immunoexpression and RAS immunoexpression on the survival and prognosis of lung adenocarcinoma patients are debated in the literature. Twenty-six patients, who underwent pulmonary resections between 2002 and 2007 in our clinic, and whose pathologic examinations yielded adenocarcinoma, were included in the study. EGFR and RAS expression levels were examined by immunohistochemical methods. The results were compared with the survival, stage of the disease, nodal involvement, lymphovascular invasion, and pleural invasion. Nonparametric bivariate analyses were used for statistical analyses. A significant link between EGFR immunoexpression and survival has been identified while RAS immunoexpression and survival have been proven to be irrelevant. Neither EGFR, nor RAS has displayed a significant link with the stage of the disease, nodal involvement, lymphovascular invasion, or pleural invasion. Positive EGFR immunoexpression affects survival negatively, while RAS immunoexpression has no effect on survival in lung adenocarcinoma patients.

The levels of mutant K-RAS and mutant N-RAS are rapidly reduced in a Beclin1 / ATG5 -dependent fashion by the irreversible ERBB1/2/4 inhibitor neratinib.

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Booth, Laurence; Roberts, Jane L; Poklepovic, Andrew; Kirkwood, John; Sander, Cindy; Avogadri-Connors, Francesca; Cutler, Richard E; Lalani, Alshad S; Dent, Paul

2018-02-01

The FDA approved irreversible inhibitor of ERBB1/2/4, neratinib, was recently shown to rapidly down-regulate the expression of ERBB1/2/4 as well as the levels of c-MET and mutant K-RAS via autophagic degradation. In the present studies, in a dose-dependent fashion, neratinib reduced the expression levels of mutant K-RAS or of mutant N-RAS, which was augmented in an additive to greater than additive fashion by the HDAC inhibitors sodium valproate and AR42. Neratinib could reduce PDGFRα levels in GBM cells, that was enhanced by sodium valproate. Knock down of Beclin1 or of ATG5 prevented neratinib and neratinib combined with sodium valproate / AR42 from reducing the expression of mutant N-RAS in established PDX and fresh PDX models of ovarian cancer and melanoma, respectively. Neratinib and the drug combinations caused the co-localization of mutant RAS proteins and ERBB2 with Beclin1 and cathepsin B. The drug combination activated the AMP-dependent protein kinase that was causal in enhancing HMG Co A reductase phosphorylation. Collectively, our data reinforce the concept that the irreversible ERBB1/2/4 inhibitor neratinib has the potential for use in the treatment of tumors expressing mutant RAS proteins.

Aurora-A overexpression enhances cell-aggregation of Ha-ras transformants through the MEK/ERK signaling pathway

International Nuclear Information System (INIS)

Tseng, Ya-Shih; Lee, Jenq-Chang; Huang, Chi-Ying F; Liu, Hsiao-Sheng

2009-01-01

Overexpression of Aurora-A and mutant Ras (Ras V12 ) together has been detected in human bladder cancer tissue. However, it is not clear whether this phenomenon is a general event or not. Although crosstalk between Aurora-A and Ras signaling pathways has been reported, the role of these two genes acting together in tumorigenesis remains unclear. Real-time PCR and sequence analysis were utilized to identify Ha- and Ki-ras mutation (Gly -> Val). Immunohistochemistry staining was used to measure the level of Aurora-A expression in bladder and colon cancer specimens. To reveal the effect of overexpression of the above two genes on cellular responses, mouse NIH3T3 fibroblast derived cell lines over-expressing either Ras V12 and wild-type Aurora-A (designated WT) or Ras V12 and kinase-inactivated Aurora-A (KD) were established. MTT and focus formation assays were conducted to measure proliferation rate and focus formation capability of the cells. Small interfering RNA, pharmacological inhibitors and dominant negative genes were used to dissect the signaling pathways involved. Overexpression of wild-type Aurora-A and mutation of Ras V12 were detected in human bladder and colon cancer tissues. Wild-type Aurora-A induces focus formation and aggregation of the Ras V12 transformants. Aurora-A activates Ral A and the phosphorylation of AKT as well as enhances the phosphorylation of MEK, ERK of WT cells. Finally, the Ras/MEK/ERK signaling pathway is responsible for Aurora-A induced aggregation of the Ras V12 transformants. Wild-type-Aurora-A enhances focus formation and aggregation of the Ras V12 transformants and the latter occurs through modulating the Ras/MEK/ERK signaling pathway

Targeting MET Amplification as a New Oncogenic Driver

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Kawakami, Hisato; Okamoto, Isamu; Okamoto, Wataru; Tanizaki, Junko; Nakagawa, Kazuhiko; Nishio, Kazuto

2014-01-01

Certain genetically defined cancers are dependent on a single overactive oncogene for their proliferation and survival, a phenomenon known as “oncogene addiction”. A new generation of drugs that selectively target such “driver oncogenes” manifests a clinical efficacy greater than that of conventional chemotherapy in appropriate genetically defined patients. MET is a proto-oncogene that encodes a receptor tyrosine kinase, and aberrant activation of MET signaling occurs in a subset of advanced cancers as result of various genetic alterations including gene amplification, polysomy, and gene mutation. Our preclinical studies have shown that inhibition of MET signaling either with the small-molecule MET inhibitor crizotinib or by RNA interference targeted to MET mRNA resulted in marked antitumor effects in cancer cell lines with MET amplification both in vitro and in vivo. Furthermore, patients with non-small cell lung cancer or gastric cancer positive for MET amplification have shown a pronounced clinical response to crizotinib. Accumulating preclinical and clinical evidence thus suggests that MET amplification is an “oncogenic driver” and therefore a valid target for treatment. However, the prevalence of MET amplification has not been fully determined, possibly in part because of the difficulty in evaluating gene amplification. In this review, we provide a rationale for targeting this genetic alteration in cancer therapy

Targeting MET Amplification as a New Oncogenic Driver

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Kawakami, Hisato [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-Sayama, Osaka 589-8511 (Japan); Okamoto, Isamu, E-mail: okamotoi@kokyu.med.kyushu-u.ac.jp [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-Sayama, Osaka 589-8511 (Japan); Center for Clinical and Translational Research, Kyushu University Hospital, 3-1-1 Maidashi, Higashiku, Fukuoka 812-8582 (Japan); Okamoto, Wataru [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-Sayama, Osaka 589-8511 (Japan); Division of Transrlational Research, Exploratory Oncology Research & Clinical Trial Center, National Cancer Center, 6-5-1 Kashiwanoha, Kashiwa, Chiba 277-8577 (Japan); Tanizaki, Junko [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-Sayama, Osaka 589-8511 (Japan); Lowe Center for Thoracic Oncology, Dana-Farber Cancer Institute, HIM223, 450 Brookline Avenue, Boston, MA 02215 (United States); Nakagawa, Kazuhiko [Department of Medical Oncology, Kinki University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-Sayama, Osaka 589-8511 (Japan); Nishio, Kazuto [Department of Genome Biology, Kinki University Faculty of Medicine, 377-2 Ohno-higashi, Osaka-Sayama, Osaka 589-8511 (Japan)

2014-07-22

Certain genetically defined cancers are dependent on a single overactive oncogene for their proliferation and survival, a phenomenon known as “oncogene addiction”. A new generation of drugs that selectively target such “driver oncogenes” manifests a clinical efficacy greater than that of conventional chemotherapy in appropriate genetically defined patients. MET is a proto-oncogene that encodes a receptor tyrosine kinase, and aberrant activation of MET signaling occurs in a subset of advanced cancers as result of various genetic alterations including gene amplification, polysomy, and gene mutation. Our preclinical studies have shown that inhibition of MET signaling either with the small-molecule MET inhibitor crizotinib or by RNA interference targeted to MET mRNA resulted in marked antitumor effects in cancer cell lines with MET amplification both in vitro and in vivo. Furthermore, patients with non-small cell lung cancer or gastric cancer positive for MET amplification have shown a pronounced clinical response to crizotinib. Accumulating preclinical and clinical evidence thus suggests that MET amplification is an “oncogenic driver” and therefore a valid target for treatment. However, the prevalence of MET amplification has not been fully determined, possibly in part because of the difficulty in evaluating gene amplification. In this review, we provide a rationale for targeting this genetic alteration in cancer therapy.

One-way membrane trafficking of SOS in receptor-triggered Ras activation.

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Christensen, Sune M; Tu, Hsiung-Lin; Jun, Jesse E; Alvarez, Steven; Triplet, Meredith G; Iwig, Jeffrey S; Yadav, Kamlesh K; Bar-Sagi, Dafna; Roose, Jeroen P; Groves, Jay T

2016-09-01

SOS is a key activator of the small GTPase Ras. In cells, SOS-Ras signaling is thought to be initiated predominantly by membrane recruitment of SOS via the adaptor Grb2 and balanced by rapidly reversible Grb2-SOS binding kinetics. However, SOS has multiple protein and lipid interactions that provide linkage to the membrane. In reconstituted-membrane experiments, these Grb2-independent interactions were sufficient to retain human SOS on the membrane for many minutes, during which a single SOS molecule could processively activate thousands of Ras molecules. These observations raised questions concerning how receptors maintain control of SOS in cells and how membrane-recruited SOS is ultimately released. We addressed these questions in quantitative assays of reconstituted SOS-deficient chicken B-cell signaling systems combined with single-molecule measurements in supported membranes. These studies revealed an essentially one-way trafficking process in which membrane-recruited SOS remains trapped on the membrane and continuously activates Ras until being actively removed via endocytosis.

Galectin-3 mediates cross-talk between K-Ras and Let-7c tumor suppressor microRNA.

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Ran Levy

Full Text Available BACKGROUND: Galectin-3 (Gal-3 and active (GTP-bound K-Ras contribute to the malignant phenotype of many human tumors by increasing the rate of cell proliferation, survival, and migration. These Gal-3-mediated effects result from a selective binding to K-Ras.GTP, causing increased nanoclustering in the cell membrane and leading to robust Ras signaling. Regulation of the interactions between Gal-3 and active K-Ras is not fully understood. METHODS AND FINDINGS: To gain a better understanding of what regulates the critical interactions between these two proteins, we examined the role of Gal-3 in the regulation of K-Ras by using Gal-3-knockout mouse embryonic-fibroblasts (Gal-3-/- MEFs and/or Gal-3/Gal-1 double-knockout MEFs. We found that knockout of Gal-3 induced strong downregulation (∼60% of K-Ras and K-Ras.GTP. The downregulation was somewhat more marked in the double-knockout MEFs, in which we also detected robust inhibition(∼50% of ERK and Akt activation. These additional effects are probably attributable to inhibition of the weak interactions of K-Ras.GTP with Gal-1. Re-expression of Gal-3 reversed the phenotype of the Gal-3-/- MEFs and dramatically reduced the disappearance of K-Ras in the presence of cycloheximide to the levels seen in wild-type MEFs. Furthermore, phosphorylation of Gal-3 by casein kinase-1 (CK-1 induced translocation of Gal-3 from the nucleus to the cytoplasm and the plasma membrane, leading to K-Ras stabilization accompanied by downregulation of the tumor suppressor miRNA let-7c, known to negatively control K-Ras transcription. CONCLUSIONS: Our results suggest a novel cross-talk between Gal-3-mediated downregulation of let 7c microRNA (which in turn negatively regulates K-Ras transcription and elucidates the association among Gal-3 let-7c and K-Ras transcription/translation, cellular compartmentalization and activity.

Oncogenic osteomalacia associated with soft tissue chondromyxoid fibroma

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Park, Jeong Mi E-mail: jmpark@cmc.cuk.ac.kr; Woo, Young Kyun; Kang, Moo Il; Kang, Chang Suk; Hahn, Seong Tae

2001-08-01

Oncogenic osteomalacia is a rarely described clinical entity characterized by hypophosphatemia, phosphaturia, and a low concentration of 1,25-dihydroxyvitamin D{sub 3}. It is most often associated with benign mesenchymal tumor and can be cured with surgical removal of the tumor. In this paper, we present a case of oncogenic osteomalacia caused by chondromyxoid fibroma in the soft tissue of the sole of the foot in a 56-year-old woman.

The Protective Arm of the Renin Angiotensin System (RAS)

DEFF Research Database (Denmark)

understanding of the protective side of the Renin Angiotensin System (RAS) involving angiotensin AT2 receptor, ACE2, and Ang(1-7)/Mas receptor Combines the knowledge of editors who pioneered research on the protective renin angiotensin system including; Dr. Thomas Unger, one of the founders of AT2 receptor......The Protective Arm of the Renin Angiotensin System: Functional Aspects and Therapeutic Implications is the first comprehensive publication to signal the protective role of a distinct part of the renin-angiotensin system (RAS), providing readers with early insight into a complex system which...... will become of major medical importance in the near future. Focusing on recent research, The Protective Arm of the Renin Angiotensin System presents a host of new experimental studies on specific components of the RAS, namely angiotensin AT2 receptors (AT2R), the angiotensin (1-7) peptide with its receptor...

EphB4 promotes or suppresses Ras/MEK/ERK pathway in a context-dependent manner: Implications for EphB4 as a cancer target.

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Xiao, Zhan; Carrasco, Rosa; Kinneer, Krista; Sabol, Darrin; Jallal, Bahija; Coats, Steve; Tice, David A

2012-06-01

EphB4 is a member of the Eph receptor tyrosine kinase family shown to act in neuronal guidance and mediate venal/arterial separation. In contrast to these more established roles, EphB4's function in cancer is much less clear. Here we illustrate both tumor promoting as well as suppressing roles of EphB4, by showing that its activation resulted in inhibition of the Ras/ERK pathway in endothelial cells but activation of the same pathway in MCF-7 breast cancer cells. This was true if EphB4 was stimulated with EphrinB2, its natural ligand, or an agonistic monoclonal antibody for EphB4. Correspondingly, EphB4 activation stimulated MCF7 growth while inhibiting HUVEC cell proliferation. The reason for these dramatic differences is due to functional coupling of EphB4 to different downstream effectors. Reduction of p120 RasGAP in HUVEC cells attenuated the inhibitory effect of EphB4 activation on the ERK pathway, whereas knockdown of PP2A in MCF7 cells attenuated EphB4 activation of the ERK pathway. This represents the first time a functional coupling between Eph receptor and PP2A has been demonstrated leading to activation of an oncogenic pathway. Our study illustrates the caveats and potential challenges of targeting EphB4 for cancer therapy due to the conflicting effects on cancer cell and endothelial cell compartments.

Identification and preliminary characterization of protein-cysteine farnesyltransferase

International Nuclear Information System (INIS)

Manne, V.; Roberts, D.; Tobin, A.; O'Rourke, E.; Barbacid, M.; De Virgilio, M.; Meyers, C.; Ahmed, N.; Kurz, B.; Resh, M.; Kung, Hsiang-Fu

1990-01-01

Ras proteins must be isoprenylated at a conserved cysteine residue near the carboxyl terminus in order to exert their biological activity. Previous studies indicate that an intermediate in the mevalonate pathway, most likely farnesyl pyrophosphate, is the donor of this isoprenyl group. Inhibition of mevalonate synthesis reverts the abnormal phenotypes induced by the mutant RAS2 Valendash19 gene in Saccharomyces cerevisiae and blocks the maturation of Xenopus oocytes induced by an onocogenic Ras p21 protein of human origin. These results have raised the possibility of using inhibitors of the mevalonate pathway to block the transforming properties of ras oncogenes. Unfortunately, mevalonate is a precursor of various end products essential to mammalian cells, such as dolichols, ubiquinones, heme A, and cholesterol. In this study, the authors describe an enzymatic activity(ies) capable of catalyzing the farnesylation of unprocessed Ras p21 proteins in vitro at the correct (Cys-186) residue. Gel filtration analysis of a partially purified preparation of protein farnesyltransferase revealed two peaks of activity at 250-350 kDa and 80-130 kDa. Availability of an in vitro protein farnesyltransferase assay should be useful in screening for potential inhibitors of ras oncogene function that will not interfere with other aspects of the mevalonate pathway

Involvement of H- and N-Ras isoforms in transforming growth factor-β1-induced proliferation and in collagen and fibronectin synthesis

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Martinez-Salgado, Carlos; Fuentes-Calvo, Isabel; Garcia-Cenador, Begona; Santos, Eugenio; Lopez-Novoa, Jose M.

2006-01-01

Transforming growth factor β1 (TGF-β1) has a relevant role in the origin and maintenance of glomerulosclerosis and tubule-interstitial fibrosis. TGF-β and Ras signaling pathways are closely related: TGF-β1 overcomes Ras mitogenic effects and Ras counteracts TGF-β signaling. Tubule-interstitial fibrosis is associated to increases in Ras, Erk, and Akt activation in a renal fibrosis model. We study the role of N- and H-Ras isoforms, and the involvement of the Ras effectors Erk and Akt, in TGF-β1-mediated extracellular matrix (ECM) synthesis and proliferation, using embrionary fibroblasts from double knockout (KO) mice for H- and N-Ras (H-ras -/- /N-ras -/- ) isoforms and from heterozygote mice (H-ras +/- /N-ras +/- ). ECM synthesis is increased in basal conditions in H-ras -/- /N-ras -/- fibroblasts, this increase being higher after stimulation with TGF-β1. TGF-β1-induced fibroblast proliferation is smaller in H-ras -/- /N-ras -/- than in H-ras +/- /N-ras +/- fibroblasts. Erk activation is decreased in H-ras -/- /N-ras -/- fibroblasts; inhibition of Erk activation reduces fibroblast proliferation. Akt activation is higher in double KO fibroblasts than in heterozygotes; inhibition of Akt activation also inhibits ECM synthesis. We suggest that H- and N-Ras isoforms downregulate ECM synthesis, and mediate proliferation, in part through MEK/Erk activation. PI3K-Akt pathway activation may be involved in the increase in ECM synthesis observed in the absence of H- and N-Ras

Epigenetic Pathways of Oncogenic Viruses: Therapeutic Promises.

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El-Araby, Amr M; Fouad, Abdelrahman A; Hanbal, Amr M; Abdelwahab, Sara M; Qassem, Omar M; El-Araby, Moustafa E

2016-02-01

Cancerous transformation comprises different events that are both genetic and epigenetic. The ultimate goal for such events is to maintain cell survival and proliferation. This transformation occurs as a consequence of different features such as environmental and genetic factors, as well as some types of infection. Many viral infections are considered to be causative agents of a number of different malignancies. To convert normal cells into cancerous cells, oncogenic viruses must function at the epigenetic level to communicate with their host cells. Oncogenic viruses encode certain epigenetic factors that lead to the immortality and proliferation of infected cells. The epigenetic effectors produced by oncogenic viruses constitute appealing targets to prevent and treat malignant diseases caused by these viruses. In this review, we highlight the importance of epigenetic reprogramming for virus-induced oncogenesis, with special emphasis on viral epigenetic oncoproteins as therapeutic targets. The discovery of molecular components that target epigenetic pathways, especially viral factors, is also discussed. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Dietary turmeric modulates DMBA-induced p21ras, MAP kinases and AP-1/NF-κB pathway to alter cellular responses during hamster buccal pouch carcinogenesis

International Nuclear Information System (INIS)

Garg, Rachana; Ingle, Arvind; Maru, Girish

2008-01-01

The chemopreventive efficacy of turmeric has been established in experimental systems. However, its mechanism(s) of action are not fully elucidated in vivo. The present study investigates the mechanism of turmeric-mediated chemoprevention in 7,12-dimethylbenz(a)anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis at 2, 4, 6, 10 and 12 weeks. Dietary turmeric (1%) led to decrease in DMBA-induced tumor burden and multiplicity, and enhanced the latency period in parallel, to its modulatory effects on oncogene products and various cellular responses during HBP tumorigenesis. DMBA-induced expression of ras oncogene product, p21 and downstream target, the mitogen-activated protein kinases were significantly decreased by turmeric during HBP carcinogenesis. Turmeric also diminished the DMBA-induced mRNA expression of proto-oncogenes (c-jun, c-fos) and NF-κB, leading to decreased protein levels and in further attenuation of DMBA-induced AP-1/NF-κB DNA-binding in the buccal pouch nuclear extracts. Besides, buccal pouch of hamsters receiving turmeric diet showed significant alterations in DMBA-induced effects: (a) decrease in cell proliferation (diminished PCNA and Bcl2 expression), (b) enhanced apoptosis (increased expression of Bax, caspase-3 and apoptotic index), (c) decrease in inflammation (levels of Cox-2, the downstream target of AP-1/NF-κB, and PGE2) and (d) aberrant expression of differentiation markers, the cytokeratins (1, 5, 8, and 18). Together, the protective effects of dietary turmeric converge on augmenting apoptosis of the initiated cells and decreasing cell proliferation in DMBA-treated animals, which in turn, is reflected in decreased tumor burden, multiplicity and enhanced latency period. Some of these biomarkers are likely to be helpful in monitoring clinical trials and evaluating drug effect measurements

Pressure modulation of Ras-membrane interactions and intervesicle transfer.

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Kapoor, Shobhna; Werkmüller, Alexander; Goody, Roger S; Waldmann, Herbert; Winter, Roland

2013-04-24

Proteins attached to the plasma membrane frequently encounter mechanical stresses, including high hydrostatic pressure (HHP) stress. Signaling pathways involving membrane-associated small GTPases (e.g., Ras) have been identified as critical loci for pressure perturbation. However, the impact of mechanical stimuli on biological outputs is still largely terra incognita. The present study explores the effect of HHP on the membrane association, dissociation, and intervesicle transfer process of N-Ras by using a FRET-based assay to obtain the kinetic parameters and volumetric properties along the reaction path of these processes. Notably, membrane association is fostered upon pressurization. Conversely, depending on the nature and lateral organization of the lipid membrane, acceleration or retardation is observed for the dissociation step. In addition, HHP can be inferred as a positive regulator of N-Ras clustering, in particular in heterogeneous membranes. The susceptibility of membrane interaction to pressure raises the idea of a role of lipidated signaling molecules as mechanosensors, transducing mechanical stimuli to chemical signals by regulating their membrane binding and dissociation. Finally, our results provide first insights into the influence of pressure on membrane-associated Ras-controlled signaling events in organisms living under extreme environmental conditions such as those that are encountered in the deep sea and sub-seafloor environments, where pressures reach the kilobar (100 MPa) range.

IQGAP1 is an oncogenic target in canine melanoma.

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Becky H Lee

Full Text Available Canine oral mucosal melanoma is an aggressive malignant neoplasm and is characterized by local infiltration and a high metastatic potential. The disease progression is similar to that of human oral melanomas. Whereas human cutaneous melanoma is primarily driven by activating mutations in Braf (60% or Nras (20%, human mucosal melanoma harbors these mutations much less frequently. This makes therapeutic targeting and research modeling of the oral form potentially different from that of the cutaneous form in humans. Similarly, research has found only rare Nras mutations and no activating Braf mutations in canine oral melanomas, but they are still reliant on MAPK signaling. IQGAP1 is a signaling scaffold that regulates oncogenic ERK1/2 MAPK signaling in human Ras- and Raf- driven cancers, including melanomas. To investigate whether IQGAP1 is a potential target in canine melanoma, we examined the expression and localization of IQGAP1 in primary canine melanomas and canine oral melanoma cell lines obtained from the University of California-Davis. Using CRISPR/Cas9 knockout of IQGAP1, we examined effects on downstream ERK1/2 pathway activity and assayed proliferation of cell lines when treated with a peptide that blocks the interaction between IQGAP1 and ERK1/2. We observed that canine IQGAP1 is expressed and localizes to a similar extent in both human and canine melanoma by qPCR, Western blot, and immunofluorescence. Deletion of IQGAP1 reduces MAPK pathway activation in cell lines, similar to effects seen in human BrafV600E cell lines. Additionally, we demonstrated reduced proliferation when these cells are treated with a blocking peptide in vitro.

Promoter trans-activation of protooncogenes c-fos and c-myc, but not c-Ha-ras, by products of adenovirus early region 1A

International Nuclear Information System (INIS)

Sassone-Corsi, P.; Borrelli, E.

1987-01-01

The E1A (early region 1A) oncogene products of adenovirus type 2 trans-activate the other early viral transcription units, as well as some cellular promoters. Using a short-term cotransfection assay in murine NIH 3T3 fibroblasts, we show that c-fos and c-myc promoter activities are stimulated by the E1A proteins, whereas c-Ha-ras transcription is not affected. The product of E1A 13S mRNA is responsible for the trans-activation, whereas the 12S mRNA product has no effect. Analysis of the c-fos promoter sequences required for the E1A stimulation shows that responsive sequences are located between positions -402 and -240 upstream of the transcription initiation site. This same region also contains the c-fos serum-responsive element. Furthermore, transcription of the endogenous c-fos gene in HeLa cells is increased after E1A transfection

Evidence implicating the Ras pathway in multiple CD28 costimulatory functions in CD4+ T cells.

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Sujit V Janardhan

Full Text Available CD28 costimulation is a critical event in the full activation of CD4(+ T cells that augments cytokine gene transcription, promotes cytokine mRNA stability, prevents induction of anergy, increases cellular metabolism, and increases cell survival. However, despite extensive biochemical analysis of the signaling events downstream of CD28, molecular pathways sufficient to functionally replace the diverse aspects of CD28-mediated costimulation in normal T cells have not been identified. Ras/MAPK signaling is a critical pathway downstream of T cell receptor stimulation, but its role in CD28-mediated costimulation has been controversial. We observed that physiologic CD28 costimulation caused a relocalization of the RasGEF RasGRP to the T cell-APC interface by confocal microscopy. In whole cell biochemical analysis, CD28 cross-linking with either anti-CD28 antibody or B7.1-Ig augmented TCR-induced Ras activation. To determine whether Ras signaling was sufficient to functionally mimic CD28 costimulation, we utilized an adenoviral vector encoding constitutively active H-Ras (61L to transduce normal, Coxsackie-Adenovirus Receptor (CAR transgenic CD4(+ T cells. Like costimulation via CD28, active Ras induced AKT, JNK and ERK phosphorylation. In addition, constitutive Ras signaling mimicked the ability of CD28 to costimulate IL-2 protein secretion, prevent anergy induction, increase glucose uptake, and promote cell survival. Importantly, we also found that active Ras mimicked the mechanism by which CD28 costimulates IL-2 production: by increasing IL-2 gene transcription, and promoting IL-2 mRNA stability. Finally, active Ras was able to induce IL-2 production when combined with ionomycin stimulation in a MEK-1-dependent fashion. Our results are consistent with a central role for Ras signaling in CD28-mediated costimulation.

Assessment of the chemosensitizing activity of TAT-RasGAP317-326 in childhood cancers.

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Nadja Chevalier

Full Text Available Although current anti-cancer protocols are reasonably effective, treatment-associated long-term side effects, induced by lack of specificity of the anti-cancer procedures, remain a challenging problem in pediatric oncology. TAT-RasGAP317-326 is a RasGAP-derived cell-permeable peptide that acts as a sensitizer to various anti-cancer treatments in adult tumor cells. In the present study, we assessed the effect of TAT-RasGAP317-326 in several childhood cancer cell lines. The RasGAP-derived peptide-induced cell death was analyzed in several neuroblastoma, Ewing sarcoma and leukemia cell lines (as well as in normal lymphocytes. Cell death was evaluated using flow cytometry methods in the absence or in the presence of the peptide in combination with various genotoxins used in the clinics (4-hydroperoxycyclophosphamide, etoposide, vincristine and doxorubicin. All tested pediatric tumors, in response to at least one genotoxin, were sensitized by TAT-RasGAP317-326. The RasGAP-derived peptide did not increase cell death of normal lymphocytes, alone or in combination with the majority of the tested chemotherapies. Consequently, TAT-RasGAP317-326 may benefit children with tumors by increasing the efficacy of anti-cancer therapies notably by allowing reductions in anti-cancer drug dosage and the associated drug-induced side effects.

Lead identification for the K-Ras protein: virtual screening and combinatorial fragment-based approaches

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Pathan AAK

2016-05-01

Full Text Available Akbar Ali Khan Pathan,1,2,* Bhavana Panthi,3,* Zahid Khan,1 Purushotham Reddy Koppula,4–6 Mohammed Saud Alanazi,1 Sachchidanand,3 Narasimha Reddy Parine,1 Mukesh Chourasia3,* 1Genome Research Chair (GRC, Department of Biochemistry, College of Science, King Saud University, 2Integrated Gulf Biosystems, Riyadh, Kingdom of Saudi Arabia; 3Department of Pharmacoinformatics, National Institute of Pharmaceutical Education and Research, Hajipur, India; 4Department of Internal Medicine, School of Medicine, 5Harry S. Truman Memorial Veterans Affairs Hospital, 6Department of Radiology, School of Medicine, Columbia, MO, USA *These authors contributed equally to this work Objective: Kirsten rat sarcoma (K-Ras protein is a member of Ras family belonging to the small guanosine triphosphatases superfamily. The members of this family share a conserved structure and biochemical properties, acting as binary molecular switches. The guanosine triphosphate-bound active K-Ras interacts with a range of effectors, resulting in the stimulation of downstream signaling pathways regulating cell proliferation, differentiation, and apoptosis. Efforts to target K-Ras have been unsuccessful until now, placing it among high-value molecules against which developing a therapy would have an enormous impact. K-Ras transduces signals when it binds to guanosine triphosphate by directly binding to downstream effector proteins, but in case of guanosine diphosphate-bound conformation, these interactions get disrupted. Methods: In the present study, we targeted the nucleotide-binding site in the “on” and “off” state conformations of the K-Ras protein to find out suitable lead compounds. A structure-based virtual screening approach has been used to screen compounds from different databases, followed by a combinatorial fragment-based approach to design the apposite lead for the K-Ras protein. Results: Interestingly, the designed compounds exhibit a binding preference for the �

The K-Ras 4A isoform promotes apoptosis but does not affect either lifespan or spontaneous tumor incidence in aging mice

International Nuclear Information System (INIS)

Plowman, Sarah J.; Arends, Mark J.; Brownstein, David G.; Luo Feijun; Devenney, Paul S.; Rose, Lorraine; Ritchie, Ann-Marie; Berry, Rachel L.; Harrison, David J.; Hooper, Martin L.; Patek, Charles E.

2006-01-01

Ras proteins function as molecular switches in signal transduction pathways, and, here, we examined the effects of the K-ras4A and 4B splice variants on cell function by comparing wild-type embryonic stem (ES) cells with K-ras tmΔ4A/tmΔ4A (exon 4A knock-out) ES cells which express K-ras4B only and K-ras -/- (exons 1-3 knock-out) ES cells which express neither splice variant, and intestinal epithelium from wild-type and K-ras tmΔ4A/tmΔ4A mice. RT-qPCR analysis found that K-ras4B expression was reduced in K-ras tmΔ4A/tmΔ4A ES cells but unaffected in small intestine. K-Ras deficiency did not affect ES cell growth, and K-Ras4A deficiency did not affect intestinal epithelial proliferation. K-ras tmΔ4A/tmΔ4A and K-ras -/- ES cells showed a reduced capacity for differentiation following LIF withdrawal, and K-ras -/- cells were least differentiated. K-Ras4A deficiency inhibited etoposide-induced apoptosis in ES cells and intestinal epithelial cells. However, K-ras tmΔ4A/tmΔ4A ES cells were more resistant to etoposide-induced apoptosis than K-ras -/- cells. The results indicate that (1) K-Ras4A promotes apoptosis while K-Ras4B inhibits it, and (2) K-Ras4B, and possibly K-Ras4A, promotes differentiation. The findings raise the possibility that alteration of the K-Ras4A/4B isoform ratio modulates tumorigenesis by differentially affecting stem cell survival and/or differentiation. However, K-Ras4A deficiency did not affect life expectancy or spontaneous overall tumor incidence in aging mice

Resistance and resilience of small-scale recirculating aquaculture systems (RAS) with or without algae to pH perturbation

Science.gov (United States)

Giatsis, Christos; Md Yusoff, Fatimah; Verreth, Johan; Verdegem, Marc

2018-01-01

The experimental set-up of this study mimicked recirculating aquaculture systems (RAS) where water quality parameters such as dissolved oxygen, pH, temperature, and turbidity were controlled and wastes produced by fish and feeding were converted to inorganic forms. A key process in the RAS was the conversion of ammonia to nitrite and nitrite to nitrate through nitrification. It was hypothesized that algae inclusion in RAS would improve the ammonia removal from the water; thereby improving RAS water quality and stability. To test this hypothesis, the stability of the microbiota community composition in a freshwater RAS with (RAS+A) or without algae (RAS-A) was challenged by introducing an acute pH drop (from pH 7 to 4 during three hours) to the system. Stigeoclonium nanum, a periphytic freshwater microalga was used in this study. No significant effect of the algae presence was found on the resistance to the acute pH drop on ammonia conversion to nitrite and nitrite conversion to nitrate. Also the resilience of the ammonia conversion to the pH drop disruption was not affected by the addition of algae. This could be due to the low biomass of algae achieved in the RAS. However, with regard to the conversion step of nitrite to nitrate, RAS+A was significantly more resilient than RAS-A. In terms of overall bacterial communities, the composition and predictive function of the bacterial communities was significantly different between RAS+A and RAS-A. PMID:29659617

Role of Notch Signaling in Human Breast Cancer Pathogenesis

Science.gov (United States)

2006-11-01

transform HMLE cells. Similarly, overexpression of ErbB2, a receptor tyrosine kinase upstream of Ras normally found overexpressed in many breast cancers ...Assess Notch-Ras cooperation in breast cancers in vivo: Since the major observation in this project has been the cooperation of Notch and Ras in HMLE ...metastasis. The in vitro cooperation between Notch and Ras in HMLE cells is mimicked in naturally arising breast cancers in vivo. Further dissection of the

Resistance to experimental tumorigenesis in cells of a long-lived mammal, the naked mole-rat (Heterocephalus glaber).

Science.gov (United States)

Liang, Sitai; Mele, James; Wu, Yuehong; Buffenstein, Rochelle; Hornsby, Peter J

2010-08-01

The naked mole-rat (NMR, Heterocephalus glaber) is a long-lived mammal in which spontaneous cancer has not been observed. To investigate possible mechanisms for cancer resistance in this species, we studied the properties of skin fibroblasts from the NMR following transduction with oncogenes that cause cells of other mammalian species to form malignant tumors. Naked mole-rat fibroblasts were transduced with a retrovirus encoding SV40 large T antigen and oncogenic Ras(G12V). Following transplantation of transduced cells into immunodeficient mice, cells rapidly entered crisis, as evidenced by the presence of anaphase bridges, giant cells with enlarged nuclei, multinucleated cells, and cells with large number of chromosomes or abnormal chromatin material. In contrast, similarly transduced mouse and rat fibroblasts formed tumors that grew rapidly without crisis. Crisis was also observed after > 40 population doublings in SV40 TAg/Ras-expressing NMR cells in culture. Crisis in culture was prevented by additional infection of the cells with a retrovirus encoding hTERT (telomerase reverse transcriptase). SV40 TAg/Ras/hTERT-expressing NMR cells formed tumors that grew rapidly in immunodeficient mice without evidence of crisis. Crisis could also be induced in SV40 TAg/Ras-expressing NMR cells by loss of anchorage, but after hTERT transduction, cells were able to proliferate normally following loss of anchorage. Thus, rapid crisis is a response of oncogene-expressing NMR cells to growth in an in vivo environment, which requires anchorage independence, and hTERT permits cells to avoid crisis and to achieve malignant tumor growth. The unique reaction of NMR cells to oncogene expression may form part of the cancer resistance of this species.

RAS in the central nervous system: Potential role in neuropsychiatric disorders.

Science.gov (United States)

Rocha, Natalia Pessoa; Simões e Silva, Ana Cristina; Prestes, Thiago Ruiz Rodrigues; Feracin, Victor; Machado, Caroline Amaral; Ferreira, Rodrigo Novaes; Teixeira, Antonio Lucio; de Miranda, Aline Silva

2018-02-25

The Renin-Angiotensin System (RAS) is a key regulator of cardiovascular and renal homeostasis, but also plays important roles in mediating physiological functions in the central nervous system (CNS). The effects of the RAS were classically described as mediated by angiotensin (Ang) II via angiotensin type 1 (AT1) receptors. However, another arm of the RAS formed by the angiotensin converting enzyme 2 (ACE2), Ang-(1-7) and the Mas receptor has been a matter of investigation due to its important physiological roles, usually counterbalancing the classical effects exerted by Ang II. We aim to provide an overview of effects elicited by the RAS, especially Ang-(1-7), in the brain. We also aim to discuss the therapeutic potential for neuropsychiatric disorders of the modulation of RAS. We carried out an extensive literature search in PubMed central. Within the brain, Ang-(1-7) contributes to the regulation of blood pressure by acting at regions that control cardiovascular functions. In contrast with Ang II, Ang-(1-7) improves baroreflex sensitivity and plays an inhibitory role in hypothalamic noradrenergic neurotransmission. Ang-(1-7) not only exerts effects related to blood pressure regulation, but also acts as a neuroprotective component of the RAS, for instance, by reducing cerebral infarct size, inflammation, oxidative stress and neuronal apoptosis. Pre-clinical evidence supports a relevant role for ACE2/Ang-(1-7)/Mas receptor axis in several neuropsychiatric conditions, including stress-related and mood disorders, cerebrovascular ischemic and haemorrhagic lesions and neurodegenerative diseases. However, very few data are available regarding the ACE2/Ang-(1-7)/Mas receptor axis in human CNS. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Safe use of NSAIDs and RAS-inhibitors at Agogo Presbyterian Hospital, Ghana

NARCIS (Netherlands)

Meulendijks, L.G.; Adomako, E.A.; Appiah, E.B.; Kramers, C.

2016-01-01

BACKGROUND: Preventable adverse events of medication are an important cause of hospital admissions in the developed world, in which non-steroidal anti-inflammatory drugs (NSAIDs) and renin-angiotensin system (RAS-) inhibitors are frequently involved. NSAIDs and RAS-inhibitors are also often used in

Molecular analysis of radiation-induced experimental tumors in mice

International Nuclear Information System (INIS)

Niwa, O.; Muto, M.; Suzuki, F.

1992-01-01

Molecular analysis was made on mouse tumors induced by radiation and chemicals. Expression of oncogenes was studied in 12 types of 178 mouse tumors. Southern blotting was done on tumors in which overexpression of oncogenes was noted. Amplification of the myc oncogene was found in chemically induced sarcomas, but not those induced by radiations. Radiogenic thymomas were studied in detail. These thymomas were induced in two different ways. The first was thymomas induced by direct irradiation of F1 mice between C57BL/6NxC3H/He. Southern analysis of DNA revealed deletion of specific minisatellite bands in these tumors. DNA from directly induced thymomas induced focus formation when transfected into normal Golden hamster cells. The mouse K-ras oncogene was detected in these transformants. The second type of thymomas was induced by X-irradiation of thymectomized B10.thy1.2 mice in which normal thymus from congenic B10,thy1.1. mice was grafted. Thymomas of the donor origin was analysed by transfection and the transformants by DNA from those indirectly induced thymomas did not contain activated ras oncogenes. (author)

Analisis Pemasaran Ayam Ras Pedaging di Pasar Tradisional Kota Medan

OpenAIRE

Fachri, Yusrizal

2017-01-01

130306044 YUSRIZAL FACHRI, 2017. “Analisis Pemasaran Ayam Ras Pedaging di Pasar Tradisional Kota Medan”. Dibimbing oleh ARMYN HAKIM DAULAY dan NEVY DIANA HANAFI. Penelitian ini bertujuan untuk mengidentifikasi karakteristik lembaga pemasaran, bentuk saluran, fungsi pemasaran, dan menganalisis nilai tambah di lembaga-lembaga pemasaran ayam ras pedaging di pasar tradisional Kota Medan pada April sampai Mei 2017. Penelitian ini menggunakan data primer yang didapatkan dari observasi dan waw...

Molecular kinetics. Ras activation by SOS: allosteric regulation by altered fluctuation dynamics.

Science.gov (United States)

Iversen, Lars; Tu, Hsiung-Lin; Lin, Wan-Chen; Christensen, Sune M; Abel, Steven M; Iwig, Jeff; Wu, Hung-Jen; Gureasko, Jodi; Rhodes, Christopher; Petit, Rebecca S; Hansen, Scott D; Thill, Peter; Yu, Cheng-Han; Stamou, Dimitrios; Chakraborty, Arup K; Kuriyan, John; Groves, Jay T

2014-07-04

Activation of the small guanosine triphosphatase H-Ras by the exchange factor Son of Sevenless (SOS) is an important hub for signal transduction. Multiple layers of regulation, through protein and membrane interactions, govern activity of SOS. We characterized the specific activity of individual SOS molecules catalyzing nucleotide exchange in H-Ras. Single-molecule kinetic traces revealed that SOS samples a broad distribution of turnover rates through stochastic fluctuations between distinct, long-lived (more than 100 seconds), functional states. The expected allosteric activation of SOS by Ras-guanosine triphosphate (GTP) was conspicuously absent in the mean rate. However, fluctuations into highly active states were modulated by Ras-GTP. This reveals a mechanism in which functional output may be determined by the dynamical spectrum of rates sampled by a small number of enzymes, rather than the ensemble average. Copyright © 2014, American Association for the Advancement of Science.

Transformation and oncogenicity by Adenoviruses

NARCIS (Netherlands)

Bernards, R.A.; Eb, A.J. van der

1984-01-01

Adenoviruses have attracted considerable attention since it was discovered by TRENTIN et all. and HUEBNER et al. that certain species (formerly called serotypes) are oncogenic when injected into newborn hamsters. Since then, adenoviruses have been used extensively as a model for studies on tumor

Targeting oncogenic Myc as a strategy for cancer treatment.

Science.gov (United States)

Chen, Hui; Liu, Hudan; Qing, Guoliang

2018-01-01

The MYC family oncogene is deregulated in >50% of human cancers, and this deregulation is frequently associated with poor prognosis and unfavorable patient survival. Myc has a central role in almost every aspect of the oncogenic process, orchestrating proliferation, apoptosis, differentiation, and metabolism. Although Myc inhibition would be a powerful approach for the treatment of many types of cancers, direct targeting of Myc has been a challenge for decades owing to its "undruggable" protein structure. Hence, alternatives to Myc blockade have been widely explored to achieve desirable anti-tumor effects, including Myc/Max complex disruption, MYC transcription and/or translation inhibition, and Myc destabilization as well as the synthetic lethality associated with Myc overexpression. In this review, we summarize the latest advances in targeting oncogenic Myc, particularly for cancer therapeutic purposes.

Modulating factors in the expression of radiation-induced oncogenic transformation

International Nuclear Information System (INIS)

Hall, E.J.; Hei, T.K.

1990-01-01

Many assays for oncogenic transformation have been developed ranging from those in established rodent cell lines where morphological alteration is scored, to those in human cells growing in nude mice where tumor invasiveness is scored. In general, systems that are most quantitaive are also the least relevant in terms of human carcinogenesis and human risk estimation. The development of cell culture systems has made it possible to assess at the cellular level the oncogenic potential of a variety of chemical, physical and viral agents. Cell culture systems afford the opportunity to identify factors and conditions that may prevent or enhance cellular transformation by radiation and chemicals. Permissive and protective factors in radiation-induced transformation include thyroid hormone and the tumor promoter TPA that increase the transformation incidence for a given dose of radiation, and retinoids, selenium, vitamin E, and 5-aminobenzamide that inhibit the expression of transformation. Densely ionizing α-particles, similar to those emitted by radon daughters, are highly effective in inducing transformations and appear to interact in a supra-additive fashion with asbestos fibers. The activation of a known dominant oncogene has not yet been demonstrated in radiation-induced oncogenic transformation. The most likely mechanism for radiation activation of an oncogene would be via the production of a chromosomal translocation. Radiation also efficiently induces deletions and may thus lead to the loss of a suppressor gene

K-Ras and mitochondria: Dangerous liaisons

Czech Academy of Sciences Publication Activity Database

Neužil, Jiří; Rohlena, Jakub; Dong, L. F.

2012-01-01

Roč. 22, č. 2 (2012), s. 285-287 ISSN 1001-0602 Institutional research plan: CEZ:AV0Z50520701 Keywords : Cancer * generation * oncogene Subject RIV: EA - Cell Biology Impact factor: 10.526, year: 2012

Demonstration of pb-PSHA with Ras-Elhekma earthquake, Egypt

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Elsayed Fergany

2017-06-01

Full Text Available The main goal of this work is to: (1 argue for the importance of a physically-based probabilistic seismic hazard analysis (pb-PSHA methodology and show examples to support the argument from recent events, (2 demonstrate the methodology with the ground motion simulations of May 28, 1998, Mw = 5.5 Ras-Elhekma earthquake, north Egypt. The boundaries for the possible rupture parameters that may have been identified prior to the 1998 Ras-Elhekma earthquake were estimated. A range of simulated ground-motions for the Ras-Elhekma earthquake was “predicted” for frequency 0.5–25 Hz at three sites, where the large earthquake was recorded, with average epicentral distances of 220 km. The best rupture model of the 1998 Ras-Elhekma earthquake was identified by calculated the goodness of fit between observed and synthesized records at sites FYM, HAG, and KOT. We used the best rupture scenario of the 1998 earthquake to synthesize the ground motions at interested sites where the main shock was not recorded. Based on the good fit of simulated and observed seismograms, we concluded that this methodology can provide realistic ground motion of an earthquake and highly recommended for engineering purposes in advance or foregoing large earthquakes at non record sites. We propose that there is a need for this methodology for good-representing the true hazard with reducing uncertainties.

Specific repression of mutant K-RAS by 10-23 DNAzyme: Sensitizing cancer cell to anti-cancer therapies

International Nuclear Information System (INIS)

Yu, S.-H.; Wang, T.-H.; Au, L.-C.

2009-01-01

Point mutations of the Ras family are frequently found in human cancers at a prevalence rate of 30%. The most common mutation K-Ras(G12V), required for tumor proliferation, survival, and metastasis due to its constitutively active GTPase activity, has provided an ideal target for cancer therapy. 10-23 DNAzyme, an oligodeoxyribonucleotide-based ribonuclease consisting of a 15-nucleotide catalytical domain flanked by two target-specific complementary arms, has been shown to effectively cleave the target mRNA at purine-pyrimidine dinucleotide. Taking advantage of this specific property, 10-23 DNAzyme was designed to cleave mRNA of K-Ras(G12V)(GGU → GUU) at the GU dinucleotide while left the wild-type (WT) K-Ras mRNA intact. The K-Ras(G12V)-specific 10-23 DNAzyme was able to reduce K-Ras(G12V) at both mRNA and protein levels in SW480 cell carrying homozygous K-Ras(G12V). No effect was observed on the WT K-Ras in HEK cells. Although K-Ras(G12V)-specific DNAzymes alone did not inhibit proliferation of SW480 or HEK cells, pre-treatment of this DNAzyme sensitized the K-Ras(G12V) mutant cells to anti-cancer agents such as doxorubicin and radiation. These results offer a potential of using allele-specific 10-23 DNAzyme in combination with other cancer therapies to achieve better effectiveness on cancer treatment.

Ras activation by SOS: Allosteric regulation by altered fluctuation dynamics

Science.gov (United States)

Iversen, Lars; Tu, Hsiung-Lin; Lin, Wan-Chen; Christensen, Sune M.; Abel, Steven M.; Iwig, Jeff; Wu, Hung-Jen; Gureasko, Jodi; Rhodes, Christopher; Petit, Rebecca S.; Hansen, Scott D.; Thill, Peter; Yu, Cheng-Han; Stamou, Dimitrios; Chakraborty, Arup K.; Kuriyan, John; Groves, Jay T.

2014-01-01

Activation of the small guanosine triphosphatase H-Ras by the exchange factor Son of Sevenless (SOS) is an important hub for signal transduction. Multiple layers of regulation, through protein and membrane interactions, govern activity of SOS. We characterized the specific activity of individual SOS molecules catalyzing nucleotide exchange in H-Ras. Single-molecule kinetic traces revealed that SOS samples a broad distribution of turnover rates through stochastic fluctuations between distinct, long-lived (more than 100 seconds), functional states. The expected allosteric activation of SOS by Ras–guanosine triphosphate (GTP) was conspicuously absent in the mean rate. However, fluctuations into highly active states were modulated by Ras-GTP. This reveals a mechanism in which functional output may be determined by the dynamical spectrum of rates sampled by a small number of enzymes, rather than the ensemble average. PMID:24994643

Molecular analysis of p53 and K-ras in lung carcinomas of coal miners

Energy Technology Data Exchange (ETDEWEB)

Sarkar, F.H.; Li, Y.W.; Vallyathan, V. [Wayne State University, Detroit, MI (United States). School of Medicine, Dept. of Pathology

2001-10-01

Thirty-three cases of non-small cell lung cancers (NSCLC) from the archives of National Coal Workers' Autopsy Study were studied for mutational alterations in p53 and K-ras using PCR-SSCP, DNA sequencing and PCR-oligonucleotide probe hybridization techniques. Mutations of the p53 were observed in 4 smokers (19%) and one in a never smoker (8%). Two polymorphisms in smokers were detected at codon 213, a common site for sequence variation. Among the smokers the p53 mutations were in the heavy smokers. In never smokers there was only a single p53 mutation and two K-ras mutations. In never smokers the frequency of K-ras mutations was similar (17%) in smokers, but one never smoker had two K-ras mutations. Mutations of p53 were more frequent in adenocarcinomas (27%) and they were AT-GC transitions. There were two large cell undifferentiated carcinomas with p53 mutation and one with a K-ras mutation. Two of the 16 squamous cell carcinomas were positive for p53 mutation, while no K-ras mutations were found in this group. The results of these preliminary studies indicate a moderately different mutational spectrum of p53 and K-ras in coal miners independent of cigarette smoking. The mutational spectrum observed in this study of coal miners with heavy cigarette smoking history suggest a protective effect of coal mine dust in preventing abnormal mutations induced by chemical carcinogens in cigarette smoke or reactive oxygen species.

Oncogenic transformation with radiation and chemicals: review

International Nuclear Information System (INIS)

Hall, E.J.; Hei, T.K.

1985-01-01

Quantitative in vitro assay systems for oncogenic transformation are a powerful research tool. They may be based on short-term cultures of hamster embryo cells, or established cell lines of mouse origin. While X-ray-induced transformation of human cells has been demonstrated, it has proved difficult to develop quantitative assay systems based on cells of human origin. The presently available quantitative assays have two quite distinct basic uses. First, they may be useful to accumulate data which is essentially pragmatic in nature. For example, they may be used to compare and contrast the oncogenic potential of chemotherapeutic agents or hypoxic cell sensitizers used or proposed in the clinic. They may be used to identify compounds that inhibit or suppress the transformation incidence resulting from known oncogenic agents, or they may be used to demonstrate the interaction between two different agents, such as radiation and asbestos. Second, they may prove to be invaluable in the study of the basic mechanisms of carcinogenesis, inasmuch as they represent models of tumourigenesis in which the various steps can be manipulated and modified more readily and in a controlled way. (author)

The levels of mutant K-RAS and mutant N-RAS are rapidly reduced in a Beclin1 / ATG5 -dependent fashion by the irreversible ERBB1/2/4 inhibitor neratinib

OpenAIRE

Booth, Laurence; Roberts, Jane L.; Poklepovic, Andrew; Kirkwood, John; Sander, Cindy; Avogadri-Connors, Francesca; Cutler Jr, Richard E.; Lalani, Alshad S.; Dent, Paul

2017-01-01

ABSTRACT The FDA approved irreversible inhibitor of ERBB1/2/4, neratinib, was recently shown to rapidly down-regulate the expression of ERBB1/2/4 as well as the levels of c-MET and mutant K-RAS via autophagic degradation. In the present studies, in a dose-dependent fashion, neratinib reduced the expression levels of mutant K-RAS or of mutant N-RAS, which was augmented in an additive to greater than additive fashion by the HDAC inhibitors sodium valproate and AR42. Neratinib could reduce PDGFR...

Oncogenic osteomalacia diagnosed by blood pool scintigraphy

International Nuclear Information System (INIS)

Palaniswamy, Shanmuga Sundaram; Subramanyam, Padma; Kumar, Harish

2011-01-01

Oncogenic osteomalacia is a rare metabolic bone disease characterized by phosphaturia and hypophosphatemia. Certain tumors secrete a phosphaturic factor, which results in this metabolic abnormality; this factor called as phosphatonin, is in fact a fibroblast growth factor 23 (FGF-23) involved closely in phosphate homeostasis and skeletogenesis. Complete excision of these tumors facilitates reversal of the problem. We have reported here the case of a patient who was crippled with this disease and on thorough investigation revealed an oncogenic osteomalacia with tumor focus in the right tibia. The tumor was identified as a mesenchymal tumor, i.e., hemangiopericytoma. Tumor excision alleviated patient symptoms with rapid symptomatic and biochemical improvement

Extracellular vesicle communication pathways as regulatory targets of oncogenic transformation.

Science.gov (United States)

Choi, Dongsic; Lee, Tae Hoon; Spinelli, Cristiana; Chennakrishnaiah, Shilpa; D'Asti, Esterina; Rak, Janusz

2017-07-01

Pathogenesis of human cancers bridges intracellular oncogenic driver events and their impact on intercellular communication. Among multiple mediators of this 'pathological connectivity' the role of extracellular vesicles (EVs) and their subsets (exosomes, ectosomes, oncosomes) is of particular interest for several reasons. The release of EVs from cancer cells represents a unique mechanism of regulated expulsion of bioactive molecules, a process that also mediates cell-to-cell transfer of lipids, proteins, and nucleic acids. Biological effects of these processes have been implicated in several aspects of cancer-related pathology, including tumour growth, invasion, angiogenesis, metastasis, immunity and thrombosis. Notably, the emerging evidence suggests that oncogenic mutations may impact several aspects of EV-mediated cell-cell communication including: (i) EV release rate and protein content; (ii) molecular composition of cancer EVs; (iii) the inclusion of oncogenic and mutant macromolecules in the EV cargo; (iv) EV-mediated release of genomic DNA; (v) deregulation of mechanisms responsible for EV biogenesis (vesiculome) and (vi) mechanisms of EV uptake by cancer cells. Intriguingly, EV-mediated intercellular transfer of mutant and oncogenic molecules between subpopulations of cancer cells, their indolent counterparts and stroma may exert profound biological effects that often resemble (but are not tantamount to) oncogenic transformation, including changes in cell growth, clonogenicity and angiogenic phenotype, or cause cell stress and death. However, several biological barriers likely curtail a permanent horizontal transformation of normal cells through EV-mediated mechanisms. The ongoing analysis and targeting of EV-mediated intercellular communication pathways can be viewed as a new therapeutic paradigm in cancer, while the analysis of oncogenic cargo contained in EVs released from cancer cells into biofluids is being developed for clinical use as a biomarker

Effect of Primary Tumor Location on Second- or Later-line Treatment Outcomes in Patients With RAS Wild-type Metastatic Colorectal Cancer and All Treatment Lines in Patients With RAS Mutations in Four Randomized Panitumumab Studies.

Science.gov (United States)

Boeckx, Nele; Koukakis, Reija; Op de Beeck, Ken; Rolfo, Christian; Van Camp, Guy; Siena, Salvatore; Tabernero, Josep; Douillard, Jean-Yves; André, Thierry; Peeters, Marc

2018-03-08

The primary tumor location has a prognostic impact in metastatic colorectal cancer (mCRC). We report the results from retrospective analyses assessing the effect of tumor location on prognosis and efficacy of second- and later-line panitumumab treatment in patients with RAS wild-type (WT) mCRC and on prognosis in all lines of treatment in patients with RAS mutant (MT) mCRC. RAS WT data (n = 483) from 2 randomized phase III panitumumab trials (ClinicalTrials.gov identifiers, NCT00339183 and NCT00113763) were analyzed for treatment outcomes stratified by tumor location. The second analysis assessed the effect of tumor location in RAS MT patients (n = 1205) from 4 panitumumab studies (ClinicalTrials.gov identifiers, NCT00364013, NCT00819780, NCT00339183, and NCT00113763). Primary tumors located in the cecum to transverse colon were coded as right-sided; those located from the splenic flexure to the rectum were coded as left-sided. Of all patients, the tumor location was ascertained for 83% to 88%; 71% to 77% of patients had left-sided tumors. RAS WT patients with right-sided tumors did worse for all efficacy parameters compared with those with left-sided tumors. The patients with left-sided tumors had better outcomes with panitumumab than with the comparator treatment. Because of the low patient numbers, no conclusions could be drawn for right-sided mCRC. The prognostic effect of tumor location on survival was unclear for RAS MT patients. These retrospective analyses have confirmed that RAS WT right-sided mCRC is associated with a poor prognosis, regardless of the treatment. RAS WT patients with left-sided tumors benefitted from the addition of panitumumab in second or later treatment lines. Further research is warranted to determine the optimum management of right-sided mCRC and RAS MT tumors. Copyright © 2018 Elsevier Inc. All rights reserved.

Tg.rasH2 Mice and not CByB6F1 Mice Should Be Used for 28-Day Dose Range Finding Studies Prior to 26-Week Tg.rasH2 Carcinogenicity Studies.

Science.gov (United States)

Paranjpe, Madhav G; Belich, Jessica; Vidmar, Tom J; Elbekai, Reem H; McKeon, Marie; Brown, Caren

Our recent retrospective analysis of data, collected from 29 Tg.rasH2 mouse carcinogenicity studies, determined how successful the strategy of choosing the high dose for the 26-week studies was based on the estimated maximum tolerated dose (EMTD) derived from earlier 28-day dose range finding (DRF) studies conducted in CByB6F1 mice. Our analysis demonstrated that the high doses applied at EMTD in the 26-week Tg.rasH2 studies failed to detect carcinogenic effects. To investigate why the dose selection process failed in the 26-week carcinogenicity studies, the initial body weights, terminal body weights, body weight gains, food consumption, and mortality from the first 4 weeks of 26-week studies with Tg.rasH2 mice were compared with 28-day DRF studies conducted with CByB6F1 mice. Both the 26-week and the earlier respective 28-day studies were conducted with the exact same vehicle, test article, and similar dose levels. The analysis of our results further emphasizes that the EMTD and subsequent lower doses, determined on the basis of the 28-day studies in CByB6F1 mice, may not be an accurate strategy for selecting appropriate dose levels for the 26-week carcinogenicity studies in Tg.rasH2 mice. Based on the analysis presented in this article, we propose that the Tg.rasH2 mice and not the CByB6F1 mice should be used in future DRF studies. The Tg.rasH2 mice demonstrate more toxicity than the CByB6F1 mice, possibly because of their smaller size compared to CByB6F1 mice. Also, the Tg.rasH2 males appear to be more sensitive than the female Tg.rasH2 mice.

Preoperative RAS Mutational Analysis Is of Great Value in Predicting Follicular Variant of Papillary Thyroid Carcinoma

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Tae Sook Hwang

2015-01-01

Full Text Available Follicular variant of papillary thyroid carcinoma (FVPTC, particularly the encapsulated subtype, often causes a diagnostic dilemma. We reconfirmed the molecular profiles in a large number of FVPTCs and investigated the efficacy of the preoperative mutational analysis in indeterminate thyroid nodules. BRAF V600E/K601E and RAS mutational analysis was performed on 187 FVPTCs. Of these, 132 (70.6% had a point mutation in one of the BRAF V600E (n=57, BRAF K601E (n=11, or RAS (n=64 genes. All mutations were mutually exclusive. The most common RAS mutations were at NRAS codon 61. FNA aspirates from 564 indeterminate nodules were prospectively tested for BRAF and RAS mutation and the surgical outcome was correlated with the mutational status. Fifty-seven and 47 cases were positive for BRAF and RAS mutation, respectively. Twenty-seven RAS-positive patients underwent surgery and all except one patient had FVPTC. The PPV and accuracy of RAS mutational analysis for predicting FVPTC were 96% and 84%, respectively. BRAF or RAS mutations were present in more than two-thirds of FVPTCs and these were mutually exclusive. BRAF mutational analysis followed by N, H, and KRAS codon 61 mutational analysis in indeterminate thyroid nodules would streamline the management of patients with malignancies, mostly FVPTC.

Intragenic origins due to short G1 phases underlie oncogene-induced DNA replication stress.

Science.gov (United States)

Macheret, Morgane; Halazonetis, Thanos D

2018-03-01

Oncogene-induced DNA replication stress contributes critically to the genomic instability that is present in cancer. However, elucidating how oncogenes deregulate DNA replication has been impeded by difficulty in mapping replication initiation sites on the human genome. Here, using a sensitive assay to monitor nascent DNA synthesis in early S phase, we identified thousands of replication initiation sites in cells before and after induction of the oncogenes CCNE1 and MYC. Remarkably, both oncogenes induced firing of a novel set of DNA replication origins that mapped within highly transcribed genes. These ectopic origins were normally suppressed by transcription during G1, but precocious entry into S phase, before all genic regions had been transcribed, allowed firing of origins within genes in cells with activated oncogenes. Forks from oncogene-induced origins were prone to collapse, as a result of conflicts between replication and transcription, and were associated with DNA double-stranded break formation and chromosomal rearrangement breakpoints both in our experimental system and in a large cohort of human cancers. Thus, firing of intragenic origins caused by premature S phase entry represents a mechanism of oncogene-induced DNA replication stress that is relevant for genomic instability in human cancer.

Oncogenic osteomalacia due to FGF23-expressing colon adenocarcinoma.

Science.gov (United States)

Leaf, David E; Pereira, Renata C; Bazari, Hasan; Jüppner, Harald

2013-03-01

Oncogenic osteomalacia, a paraneoplastic syndrome associated with hypophosphatemia due to increased urinary phosphate excretion, is caused by excessive synthesis and secretion of fibroblast growth factor 23 (FGF23), a phosphaturic hormone that is normally produced by osteocytes. Most cases of oncogenic osteomalacia have been associated with benign tumors of bone or soft tissue; however, whether malignant neoplasms can also produce and secrete FGF23 is currently unknown. The aim was to determine whether a malignant neoplasm could cause oncogenic osteomalacia through excessive production and secretion of FGF23. We describe an 80-year-old woman with stage IV colon adenocarcinoma who presented with severe hypophosphatemia (0.4 mg/dL; reference, 2.6-4.5 mg/dL). Fractional excretion of phosphate was 34% (reference, osteomalacia should be considered in the differential diagnosis for patients with a malignant tumor who present with hypophosphatemia.

Evaluation of K-ras and p53 expression in pancreatic adenocarcinoma using the cancer genome atlas.

Directory of Open Access Journals (Sweden)

Liming Lu

Full Text Available Genetic alterations in K-ras and p53 are thought to be critical in pancreatic cancer development and progression. However, K-ras and p53 expression in pancreatic adenocarcinoma have not been systematically examined in The Cancer Genome Atlas (TCGA Data Portal. Information regarding K-ras and p53 alterations, mRNA expression data, and protein/protein phosphorylation abundance was retrieved from The Cancer Genome Atlas (TCGA databases, and analyses were performed by the cBioPortal for Cancer Genomics. The mutual exclusivity analysis showed that events in K-ras and p53 were likely to co-occur in pancreatic adenocarcinoma (Log odds ratio = 1.599, P = 0.006. The graphical summary of the mutations showed that there were hotspots for protein activation. In the network analysis, no solid association between K-ras and p53 was observed in pancreatic adenocarcinoma. In the survival analysis, neither K-ras nor p53 were associated with both survival events. As in the data mining study in the TCGA databases, our study provides a new perspective to understand the genetic features of K-ras and p53 in pancreatic adenocarcinoma.

Lipoprotein internalisation induced by oncogenic AMPK activation is essential to maintain glioblastoma cell growth.

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Ríos, M; Foretz, M; Viollet, B; Prieto, A; Fraga, M; García-Caballero, T; Costoya, J A; Señarís, R

2014-12-01

Metabolic adaptations are essential during tumour growth to maintain the high proliferation levels exhibited by cancer cells. In this study, we examined the transformations that occurred in the lipid metabolism in astrocytic tumours, and the possible role of the fuel-sensing enzyme AMPK. Metabolic targets might help design new and effective drugs for cancer. To accomplish this objective, we studied both mice and human astrocytic tumours. We first used a mouse model of astrocytoma driven by oncogenic H-RasV12 and/or with PTEN deletion based on the common constitutive activation of the Raf/MEK/ERK and PI3K/AKT cascades in human astrocytomas. We then confirmed the results in human glioblastoma cell lines and in glioblastoma tissue samples from patients. We show that the high levels of activated AMPK, observed in astrocytic tumours, increase extracellular lipid internalisation and reduce energy expenditure by inhibiting 'de novo' fatty acid (FA) synthesis, which allows tumour cells to obtain building blocks and energy to be able to create new organelles and new cells. Our findings demonstrate that AMPK plays a crucial role in glioblastoma cell growth and suggest that blocking lipoprotein receptors could potentially be used as a plausible therapeutic approach for these and other type of tumours with high levels of AMPK. Copyright © 2014 Elsevier Ltd. All rights reserved.

Ras Umm Sidd Oxygen Isotope (delta 18O) Data for 1750 to 1995

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National Oceanic and Atmospheric Administration, Department of Commerce — Ras Umm Sidd bimonthly coral oxygen isotope data (coral core RUS-95). Notes on the data: File (Ras Umm Sidd d18O.txt.) includes columns for Year AD (bimonthly...

Oncogene-inducible organoids as a miniature platform to assess cancer characteristics

NARCIS (Netherlands)

Mizutani, Tomohiro; Tsukamoto, Yoshiyuki; Clevers, Hans

2017-01-01

Direct effects of oncogenic proteins or inhibitor treatments on signaling pathways are difficult to assess in transgenic mice. In this issue, Riemer et al. (2017. J. Cell Biol. https://doi.org/10.1083/jcb.201610058) demonstrate that oncogene-inducible organoids offer the experimental versatility of

Membrane curvature enables N-Ras lipid anchor sorting to liquid-ordered membrane phases

DEFF Research Database (Denmark)

Larsen, Jannik Bruun; Jensen, Martin Borch; Bhatia, Vikram Kjøller

2015-01-01

Trafficking and sorting of membrane-anchored Ras GTPases are regulated by partitioning between distinct membrane domains. Here, in vitro experiments and microscopic molecular theory reveal membrane curvature as a new modulator of N-Ras lipid anchor and palmitoyl chain partitioning. Membrane...

Cost-effectiveness of RAS screening before monoclonal antibodies therapy in metastatic colorectal cancer based on FIRE3 Study

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Wen, Feng; Yang, Yu; Zhang, Pengfei; Zhang, Jian; Zhou, Jing; Tang, Ruilei; Chen, Hongdou; Zheng, Hanrui; Fu, Ping; Li, Qiu

2015-01-01

The surprising results published by FIRE-3 revealed that the overall survival (OS) of RAS wild-type metastatic colorectal cancer (mCRC) patients treated with Cetuximab(Cmab) and FOLFIRI combination was prolonged to 33.1 months. The substantial increase in testing and treatment costs, however, impose a considerable health burden on patients and society. Hence the study was aimed to assess the cost-effectiveness of RAS screening before monoclonal antibodies (mAbs) therapy based on FIRE-3 study. Four groups were analyzed: group 1, patients with KRAS testing treated with Cmab and FOLFIRI; group 2, patients with RAS testing treated with Cmab and FOLFIRI; group 3, patients with KRAS testing treated with bevacizumab(Bmab) and FOLFIRI; group 4, patients with RAS testing treated with Bmab and FOLFIRI. A Markov model comprising 3 health states (progression-free survival, progressive disease and death) was built. The costs were calculated from a Chinese payer perspective, and survival was reported in quality-adjusted life-months (QALMs). Average total lifetime costs ranged from $104,682.44 (RAS-Bmab) to $136,867.44 (RAS-Cmab), while the survival gained varied from 16.88 QALMs in RAS-Bmab to 21.85 QALMs in RAS-Cmab. The cost per QALM was $6,263.86 for RAS-Cmab, $6,145.84 for KRAS-Bmab, $6,201.57 for RAS-Bmab and $6,960.70 for KRAS-Cmab respectively. The KRAS-Cmab strategy was dominated by the other 3 groups. The first-treatment cost of RAS-Cmab was the most influential one to the model. In all, the RAS screening prior to Cmab treatment in mCRC seems to be a cost-effective strategy in the time of monoclonal antibodies (mAbs) therapy with the most gained QALMs. PMID:26418570

Signal interaction of Hedgehog/GLI and epidermal growth factor receptor signaling in cancer development

International Nuclear Information System (INIS)

Eberl, M.

2012-01-01

The subject of this PhD thesis is based on the cooperation of Hedgehog (HH)/GLI with epidermal growth factor receptor (EGFR) signaling synergistically promoting oncogenic transformation and cancer growth. In previous studies we have demonstrated that the HH/GLI and EGFR signaling pathways interact synergistically resulting not only in selective induction of HH/GLI-EGFR target genes, but also in the onset of oncogenic transformation and tumor formation (Kasper, Schnidar et al. 2006; Schnidar, Eberl et al. 2009). However, the molecular key mediators acting downstream of HH/GLI and EGFR signal cooperation were largely unknown and the in vivo evidence for the therapeutic relevance of HH/GLI and EGFR signal cooperation in HH-associated cancers was lacking. During my PhD thesis I could demonstrate that the integration of EGFR and HH/GLI signaling involves activation of RAS/MEK/ERK and JUN/AP1 signaling in response to EGFR activation. Furthermore I succeeded in identifying genes, including stem cell- (SOX2, SOX9), tumor growth- (JUN, TGFA, FGF19) and metastasis-associated genes (SPP1/osteopontin, CXCR4) that showed synergistic transcriptional activation by HH/GLI-EGFR signal integration. Importantly, I could demonstrate that these genes arrange themselves within a stable interdependent signaling network, which is required for in vivo growth of basal cell carcinoma (BCC) and tumor-initiating pancreatic cancer cells. These data validate EGFR signaling as additional drug target in HH/GLI driven cancers and provide new therapeutic strategies based on combined targeting of cooperative HH/GLI-EGFR signaling and selected downstream target genes (Eberl, Klingler et al. 2012). (author) [de

Distinct roles of the RasGAP family proteins in C. elegans associative learning and memory.

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Gyurkó, M Dávid; Csermely, Péter; Sőti, Csaba; Steták, Attila

2015-10-15

The Ras GTPase activating proteins (RasGAPs) are regulators of the conserved Ras/MAPK pathway. Various roles of some of the RasGAPs in learning and memory have been reported in different model systems, yet, there is no comprehensive study to characterize all gap genes in any organism. Here, using reverse genetics and neurobehavioural tests, we studied the role of all known genes of the rasgap family in C. elegans in associative learning and memory. We demonstrated that their proteins are implicated in different parts of the learning and memory processes. We show that gap-1 contribute redundantly with gap-3 to the chemosensation of volatile compounds, gap-1 plays a major role in associative learning, while gap-2 and gap-3 are predominantly required for short- and long-term associative memory. Our results also suggest that the C. elegans Ras orthologue let-60 is involved in multiple processes during learning and memory. Thus, we show that the different classes of RasGAP proteins are all involved in cognitive function and their complex interplay ensures the proper formation and storage of novel information in C. elegans.

Disorders of dysregulated signal traffic through the RAS-MAPK pathway: phenotypic spectrum and molecular mechanisms.

Science.gov (United States)

Tartaglia, Marco; Gelb, Bruce D

2010-12-01

RAS GTPases control a major signaling network implicated in several cellular functions, including cell fate determination, proliferation, survival, differentiation, migration, and senescence. Within this network, signal flow through the RAF-MEK-ERK pathway-the first identified mitogen-associated protein kinase (MAPK) cascade-mediates early and late developmental processes controlling morphology determination, organogenesis, synaptic plasticity, and growth. Signaling through the RAS-MAPK cascade is tightly controlled; and its enhanced activation represents a well-known event in oncogenesis. Unexpectedly, in the past few years, inherited dysregulation of this pathway has been recognized as the cause underlying a group of clinically related disorders sharing facial dysmorphism, cardiac defects, reduced postnatal growth, ectodermal anomalies, variable cognitive deficits, and susceptibility to certain malignancies as major features. These disorders are caused by heterozygosity for mutations in genes encoding RAS proteins, regulators of RAS function, modulators of RAS interaction with effectors, or downstream signal transducers. Here, we provide an overview of the phenotypic spectrum associated with germline mutations perturbing RAS-MAPK signaling, the unpredicted molecular mechanisms converging toward the dysregulation of this signaling cascade, and major genotype-phenotype correlations. © 2010 New York Academy of Sciences.

Cooperation of the BTB-Zinc finger protein, Abrupt, with cytoskeletal regulators in Drosophila epithelial tumorigenesis

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Nezaket Turkel

2015-08-01

Full Text Available The deregulation of cell polarity or cytoskeletal regulators is a common occurrence in human epithelial cancers. Moreover, there is accumulating evidence in human epithelial cancer that BTB-ZF genes, such as Bcl6 and ZBTB7A, are oncogenic. From our previous studies in the vinegar fly, Drosophila melanogaster, we have identified a cooperative interaction between a mutation in the apico-basal cell polarity regulator Scribble (Scrib and overexpression of the BTB-ZF protein Abrupt (Ab. Herein, we show that co-expression of ab with actin cytoskeletal regulators, RhoGEF2 or Src64B, in the developing eye-antennal epithelial tissue results in the formation of overgrown amorphous tumours, whereas ab and DRac1 co-expression leads to non-cell autonomous overgrowth. Together with ab, these genes affect the expression of differentiation genes, resulting in tumours locked in a progenitor cell fate. Finally, we show that the expression of two mammalian genes related to ab, Bcl6 and ZBTB7A, which are oncogenes in mammalian epithelial cancers, significantly correlate with the upregulation of cytoskeletal genes or downregulation of apico-basal cell polarity neoplastic tumour suppressor genes in colorectal, lung and other human epithelial cancers. Altogether, this analysis has revealed that upregulation of cytoskeletal regulators cooperate with Abrupt in Drosophila epithelial tumorigenesis, and that high expression of human BTB-ZF genes, Bcl6 and ZBTB7A, shows significant correlations with cytoskeletal and cell polarity gene expression in specific epithelial tumour types. This highlights the need for further investigation of the cooperation between these genes in mammalian systems.

P53 suppresses expression of the 14-3-3gamma oncogene

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Qi Wenqing

2011-08-01

Full Text Available Abstract Background 14-3-3 proteins are a family of highly conserved proteins that are involved in a wide range of cellular processes. Recent evidence indicates that some of these proteins have oncogenic activity and that they may promote tumorigenesis. We previously showed that one of the 14-3-3 family members, 14-3-3gamma, is over expressed in human lung cancers and that it can induce transformation of rodent cells in vitro. Methods qRTPCR and Western blot analysis were performed to examine 14-3-3gamma expression in non-small cell lung cancers (NSCLC. Gene copy number was analyzed by qPCR. P53 mutations were detected by direct sequencing and also by western blot. CHIP and yeast one hybrid assays were used to detect p53 binding to 14-3-3gamma promoter. Results Quantitative rtPCR results showed that the expression level of 14-3-3gamma was elevated in the majority of NSCLC that we examined which was also consistent with protein expression. Further analysis of the expression pattern of 14-3-3gamma in lung tumors showed a correlation with p53 mutations suggesting that p53 might suppress 14-3-3 gamma expression. Analysis of the gamma promoter sequence revealed the presence of a p53 consensus binding motif and in vitro assays demonstrated that wild-type p53 bound to this motif when activated by ionizing radiation. Deletion of the p53 binding motif eliminated p53's ability to suppress 14-3-3gamma expression. Conclusion Increased expression of 14-3-3gamma in lung cancer coincides with loss of functional p53. Hence, we propose that 14-3-3gamma's oncogenic activities cooperate with loss of p53 to promote lung tumorigenesis.

Ras conformational switching: simulating nucleotide-dependent conformational transitions with accelerated molecular dynamics.

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Barry J Grant

2009-03-01

Full Text Available Ras mediates signaling pathways controlling cell proliferation and development by cycling between GTP- and GDP-bound active and inactive conformational states. Understanding the complete reaction path of this conformational change and its intermediary structures is critical to understanding Ras signaling. We characterize nucleotide-dependent conformational transition using multiple-barrier-crossing accelerated molecular dynamics (aMD simulations. These transitions, achieved for the first time for wild-type Ras, are impossible to observe with classical molecular dynamics (cMD simulations due to the large energetic barrier between end states. Mapping the reaction path onto a conformer plot describing the distribution of the crystallographic structures enabled identification of highly populated intermediate structures. These structures have unique switch orientations (residues 25-40 and 57-75 intermediate between GTP and GDP states, or distinct loop3 (46-49, loop7 (105-110, and alpha5 C-terminus (159-166 conformations distal from the nucleotide-binding site. In addition, these barrier-crossing trajectories predict novel nucleotide-dependent correlated motions, including correlations of alpha2 (residues 66-74 with alpha3-loop7 (93-110, loop2 (26-37 with loop10 (145-151, and loop3 (46-49 with alpha5 (152-167. The interconversion between newly identified Ras conformations revealed by this study advances our mechanistic understanding of Ras function. In addition, the pattern of correlated motions provides new evidence for a dynamic linkage between the nucleotide-binding site and the membrane interacting C-terminus critical for the signaling function of Ras. Furthermore, normal mode analysis indicates that the dominant collective motion that occurs during nucleotide-dependent conformational exchange, and captured in aMD (but absent in cMD simulations, is a low-frequency motion intrinsic to the structure.

Influence of feed ingredients on water quality parameters in RAS

DEFF Research Database (Denmark)

Pedersen, Per Bovbjerg; Pedersen, Lars-Flemming; Suhr, Karin Isabel

2011-01-01

Although feed by far is providing the major input to RAS, relatively little is published about the correlation between feed composition and the resulting water quality in such systems. In a set-up with 6 identical RAS, each consisting of a fish tank (0.5 m3), a swirl separator, a submerged...... had impact on water quality in the systems as well as on matter removed by the swirl separators. In the RAS water, phosphorous (Ptot and Pdiss) concentrations were reduced by guar gum. Organic matter content (CODdiss) in the water was also reduced. Corresponding to this, more dry matter, more COD...... to the systems for 49 consecutive days. Each week, 24h-water samples (1 sample/hour) were collected from each system. The sludge collected in the swirl separator that day was also collected. Water and sludge were subsequently analysed for nitrogen, phosphorous and organic matter content. Inclusion of guar gum...

The accumulation of substances in Recirculating Aquaculture Systems (RAS) affects embryonic and larval development in common carp Cyprinus carpio

NARCIS (Netherlands)

Martins, C.I.; Pristin, M.G.; Ende, S.S.W.; Eding, E.H.; Verreth, J.A.J.

2009-01-01

The accumulation of substances in Recirculating Aquaculture Systems (RAS) may impair the growth and welfare of fish. To test the severity of contaminants accumulated in RAS, early-life stages of fish were used. Ultrafiltered water from two Recirculating Aquaculture Systems (RAS), one RAS with a high

Identification of Novel Ovarian Cancer Oncogenes that Function by Regulating Exosome Function

Science.gov (United States)

2017-09-01

Novel Ovarian Cancer Oncogenes that Function by Regulating Exosome Function September 2017 x 1Sep2016...31Aug2017 Email: mbirrer@partners.org 6 Identification of Novel Ovarian Cancer Oncogenes that Function by Regulating Exosome Function xx

Programmed Cell-to-Cell Variability in Ras Activity Triggers Emergent Behaviors during Mammary Epithelial Morphogenesis

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Jennifer S. Liu

2012-11-01

Full Text Available Variability in signaling pathway activation between neighboring epithelial cells can arise from local differences in the microenvironment, noisy gene expression, or acquired genetic changes. To investigate the consequences of this cell-to-cell variability in signaling pathway activation on coordinated multicellular processes such as morphogenesis, we use DNA-programmed assembly to construct three-dimensional MCF10A microtissues that are mosaic for low-level expression of activated H-Ras. We find two emergent behaviors in mosaic microtissues: cells with activated H-Ras are basally extruded or lead motile multicellular protrusions that direct the collective motility of their wild-type neighbors. Remarkably, these behaviors are not observed in homogeneous microtissues in which all cells express the activated Ras protein, indicating that heterogeneity in Ras activity, rather than the total amount of Ras activity, is critical for these processes. Our results directly demonstrate that cell-to-cell variability in pathway activation within local populations of epithelial cells can drive emergent behaviors during epithelial morphogenesis.

The Oncogenic Risks of Diagnostic CT Scam Studies in Children

International Nuclear Information System (INIS)

Brent, R.

2004-01-01

Brenner et al (2001) reported that estimates of the exposure to children from CT scans indicates that the exposures are both higher than from conventional radiographic studies and higher than is necessary to obtain quality examinations. utilizing the oncogenic risk data from the RERF study in Japan, Brenner et al estimated that the oncogenic risk in this population of CT exposed children exposed each year would result in an additional 500 cases of cancer. This risk estimate is supported by the RERF epidemiological data obtained from the populations exposed in Hiroshima and Nagasaki. the increased risks associated with the increased exposure from CT scans have raised concern and stimulated discussion. Although there is little doubt about the benefits of CT scans in improving the health care of children, there is concern about the estimated oncogenic risk, especially since the frequency of CT studies has been increasing. Applying the oncogenic risks of ionizing radiation from the RERF data may not be appropriate for all types of radiation exposure for accurately predicting the incidence of cancer in exposed children because of the impact of 1) partial versus whole-body irradiation, and 2) the protraction of the exposure. Other population of children who have been exposed to radiation and whose incidence of cancer has been studied will be presented and those studies indicate that the risk of cancer is much lower or not increased at all with exposures in the diagnostic range. finally, the dramatic impact of the use of CT scans in clinical pediatric practice saves lives and improves diagnostic accuracy. Therefore, it is crucial that a scholarly evaluation of the risks and benefits should be initiated. The radiology community and the manufacturers have already initiated programs to decrease the exposure significantly. But it is essential that well-planned, retrospective and prospective epidemiology studies should be initiated to study the oncogenic risks. If you want to

Small G proteins Rac1 and Ras regulate serine/threonine protein phosphatase 5 (PP5)·extracellular signal-regulated kinase (ERK) complexes involved in the feedback regulation of Raf1.

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Mazalouskas, Matthew D; Godoy-Ruiz, Raquel; Weber, David J; Zimmer, Danna B; Honkanen, Richard E; Wadzinski, Brian E

2014-02-14

Serine/threonine protein phosphatase 5 (PP5, PPP5C) is known to interact with the chaperonin heat shock protein 90 (HSP90) and is involved in the regulation of multiple cellular signaling cascades that control diverse cellular processes, such as cell growth, differentiation, proliferation, motility, and apoptosis. Here, we identify PP5 in stable complexes with extracellular signal-regulated kinases (ERKs). Studies using mutant proteins reveal that the formation of PP5·ERK1 and PP5·ERK2 complexes partially depends on HSP90 binding to PP5 but does not require PP5 or ERK1/2 activity. However, PP5 and ERK activity regulates the phosphorylation state of Raf1 kinase, an upstream activator of ERK signaling. Whereas expression of constitutively active Rac1 promotes the assembly of PP5·ERK1/2 complexes, acute activation of ERK1/2 fails to influence the phosphatase-kinase interaction. Introduction of oncogenic HRas (HRas(V12)) has no effect on PP5-ERK1 binding but selectively decreases the interaction of PP5 with ERK2, in a manner that is independent of PP5 and MAPK/ERK kinase (MEK) activity, yet paradoxically requires ERK2 activity. Additional studies conducted with oncogenic variants of KRas4B reveal that KRas(L61), but not KRas(V12), also decreases the PP5-ERK2 interaction. The expression of wild type HRas or KRas proteins fails to reduce PP5-ERK2 binding, indicating that the effect is specific to HRas(V12) and KRas(L61) gain-of-function mutations. These findings reveal a novel, differential responsiveness of PP5-ERK1 and PP5-ERK2 interactions to select oncogenic Ras variants and also support a role for PP5·ERK complexes in regulating the feedback phosphorylation of PP5-associated Raf1.

Inhibition of the Ras-Net (Elk-3) pathway by a novel pyrazole that affects microtubules.

Science.gov (United States)

Wasylyk, Christine; Zheng, Hong; Castell, Christelle; Debussche, Laurent; Multon, Marie-Christine; Wasylyk, Bohdan

2008-03-01

Net (Elk-3/SAP-2/Erp) is a transcription factor that is phosphorylated and activated by the Ras-extracellular signal-regulated kinase (Erk) signaling pathway and is involved in wound healing, angiogenesis, and tumor growth. In a cell-based screen for small molecule inhibitors of Ras activation of Net transcriptional activity, we identified a novel pyrazole, XRP44X. XRP44X inhibits fibroblast growth factor 2 (FGF-2)-induced Net phosphorylation by the Ras-Erk signaling upstream from Ras. It also binds to the colchicine-binding site of tubulin, depolymerizes microtubules, stimulates cell membrane blebbing, and affects the morphology of the actin skeleton. Interestingly, Combretastin-A4, which produces similar effects on the cytoskeleton, also inhibits FGF-2 Ras-Net signaling. This differs from other classes of agents that target microtubules, which have either little effect (vincristine) or no effect (docetaxel and nocodazole) on the Ras-Net pathway. XRP44X inhibits various cellular properties, including cell growth, cell cycle progression, and aortal sprouting, similar to other molecules that bind to the tubulin colchicine site. XRP44X has the potentially interesting property of connecting two important pathways involved in cell transformation and may thereby represent an interesting class of molecules that could be developed for cancer treatment.

Use of Human Tissue to Assess the Oncogenic Activity of Melanoma-Associated Mutations

OpenAIRE

Chudnovsky, Yakov; Adams, Amy E.; Robbins, Paul B.; Lin, Qun; Khavari, Paul A.

2005-01-01

Multiple genetic alterations occur in melanoma, a lethal skin malignancy of increasing incidence1,2. These include mutations that activate Ras and two of its effector cascades, Raf and phosphoinositide 3-kinase (PI3K). Ras and Raf induction can occur via active N-Ras and B-Raf mutants as well as by gene amplification3–5. Activation of PI3K pathway components occurs by PTEN loss and by AKT amplification6–8. Melanomas also commonly display impairment of p16INK4A-CDK4-Rb and ARF-HDM2-p53 tumor s...

Suppression of survivin expression in glioblastoma cells by the Ras inhibitor farnesylthiosalicylic acid promotes caspase-dependent apoptosis.

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Blum, Roy; Jacob-Hirsch, Jasmine; Rechavi, Gideon; Kloog, Yoel

2006-09-01

The Ras inhibitor farnesylthiosalicylic acid (FTS) has been shown to induce apoptosis in glioblastoma multiforme, but its mechanism of action was unknown. We show that FTS or dominant-negative Ras, by deregulating extracellular signal-regulated kinase and Akt signaling, decreases survivin gene transcripts in U87 glioblastoma multiforme, leading to disappearance of survivin protein and cell death. FTS affected both Ras-controlled regulators of survivin transcription and Ras-regulated survival signals. Thus, Ras inhibition by FTS resulted in release of the survivin "brake" on apoptosis and in activation of the mitochondrial apoptotic pathway: dephosphorylation of Bad, activation of Bax, release of cytochrome c, and caspase activation. FTS-induced apoptosis of U87 cells was strongly attenuated by forced expression of survivin or by caspase inhibitors. These results show that resistance to apoptosis in glioblastoma multiforme can be abolished by a single Ras inhibitor, which targets both survivin, a critical inhibitor of apoptosis, and the intrinsic mitochondrial apoptotic machinery.

Dysregulated choline metabolism in T-cell lymphoma: role of choline kinase-α and therapeutic targeting

International Nuclear Information System (INIS)

Xiong, J; Bian, J; Wang, L; Zhou, J-Y; Wang, Y; Zhao, Y; Wu, L-L; Hu, J-J; Li, B; Chen, S-J; Yan, C; Zhao, W-L

2015-01-01

Cancer cells have distinct metabolomic profile. Metabolic enzymes regulate key oncogenic signaling pathways and have an essential role on tumor progression. Here, serum metabolomic analysis was performed in 45 patients with T-cell lymphoma (TCL) and 50 healthy volunteers. The results showed that dysregulation of choline metabolism occurred in TCL and was related to tumor cell overexpression of choline kinase-α (Chokα). In T-lymphoma cells, pharmacological and molecular silencing of Chokα significantly decreased Ras-GTP activity, AKT and ERK phosphorylation and MYC oncoprotein expression, leading to restoration of choline metabolites and induction of tumor cell apoptosis/necropotosis. In a T-lymphoma xenograft murine model, Chokα inhibitor CK37 remarkably retarded tumor growth, suppressed Ras-AKT/ERK signaling, increased lysophosphatidylcholine levels and induced in situ cell apoptosis/necropotosis. Collectively, as a regulatory gene of aberrant choline metabolism, Chokα possessed oncogenic activity and could be a potential therapeutic target in TCL, as well as other hematological malignancies with interrupted Ras signaling pathways

A view on EGFR-targeted therapies from the oncogene-addiction perspective.

Science.gov (United States)

Perez, Rolando; Crombet, Tania; de Leon, Joel; Moreno, Ernesto

2013-01-01

Tumor cell growth and survival can often be impaired by inactivating a single oncogen- a phenomenon that has been called as "oncogene addiction." It is in such scenarios that molecular targeted therapies may succeed. among known oncogenes, the epidermal growth factor receptor (EGFR) has become the target of different cancer therapies. So far, however, the clinical benefit from EGFR-targeted therapies has been rather limited. a critical review of the large amount of clinical data obtained with anti-EGFR agents, carried out from the perspective of the oncogene addiction concept, may help to understand the causes of the unsatisfactory results. In this article we intend to do such an exercise taking as basis for the analysis a few case studies of anti-EGFR agents that are currently in the clinic. There, the "EGFR addiction" phenomenon becomes apparent in high-responder patients. We further discuss how the concept of oncogene addiction needs to be interpreted on the light of emerging experimental evidences and ideas; in particular, that EGFR addiction may reflect the interconnection of several cellular pathways. In this regard we set forth several hypotheses; namely, that requirement of higher glucose uptake by hypoxic tumor cells may reinforce EGFR addiction; and that chronic use of EGFR-targeted antibodies in EGFR-addicted tumors would induce stable disease by reversing the malignant phenotype of cancer stem cells and also by sustaining an anti-tumor T cell response. Finally, we discuss possible reasons for the failure of certain combinatorial therapies involving anti-EGFR agents, arguing that some of these agents might produce either a negative or a positive trans-modulation effect on other oncogenes. It becomes evident that we need operational definitions of EGFR addiction in order to determine which patient populations may benefit from treatment with anti-EGFR drugs, and to improve the design of these therapies.

A view on EGFR-targeted therapies from the oncogene-addiction perspective

Directory of Open Access Journals (Sweden)

Rolando ePerez

2013-04-01

Full Text Available Tumor cell growth and survival can often be impaired by inactivating a single oncogen – a phenomenon that has been called as 'oncogene addiction'. It is in such scenarios that molecular targeted therapies may succeed. Among known oncogenes, the epidermal growth factor receptor (EGFR has become the target of different cancer therapies. So far, however, the clinical benefit from EGFR-targeted therapies has been rather limited. A critical review of the large amount of clinical data obtained with anti-EGFR agents, carried out from the perspective of the oncogene addiction concept, may help to understand the causes of the unsatisfactory results. In this article we intend to do such an exercise taking as basis for the analysis a few case studies of anti-EGFR agents that are currently in the clinic. There, the 'EGFR addiction' phenomenon becomes apparent in high-responder patients. We further discuss how the concept of oncogene addiction needs to be interpreted on the light of emerging experimental evidences and ideas; in particular, that EGFR addiction may reflect the interconnection of several cellular pathways. In this regard we set forth several hypotheses; namely, that requirement of higher glucose uptake by hypoxic tumor cells may reinforce EGFR addiction; and that chronic use of EGFR-targeted antibodies in EGFR-addicted tumors would induce stable disease by reversing the malignant phenotype of cancer stem cells and also by sustaining an anti-tumor T cell response. Finally, we discuss possible reasons for the failure of certain combinatorial therapies involving anti-EGFR agents, arguing that some of these agents might produce either a negative or a positive trans-modulation effect on other oncogenes. It becomes evident that we need operational definitions of EGFR addiction in order to determine which patient populations may benefit from treatment with anti-EGFR drugs, and to improve the design of these therapies.

Molecular biology III - Oncogenes and tumor suppressor genes

International Nuclear Information System (INIS)

Giaccia, Amato J.

1996-01-01

Purpose: The purpose of this course is to introduce to radiation oncologists the basic concepts of tumorigenesis, building on the information that will be presented in the first and second part of this series of lectures. Objective: Our objective is to increase the current understanding of radiation oncologists with the process of tumorigenesis, especially focusing on genes that are altered in many tumor types that are potential candidates for novel molecular strategies. As strategies to treat cancer of cancer are becoming more sophisticated, it will be important for both the practitioner and academician to develop a basic understanding of the function of cancer 'genes'. This will be the third in a series of refresher courses that are meant to address recent advances in Cancer Biology in a way that both clinicians without previous knowledge of molecular biology or experienced researchers will find interesting. The lecture will begin with a basic overview of tumorigenesis; methods of detecting chromosome/DNA alterations, approaches used to isolate oncogenes and tumor suppressor genes, and their role in cell killing by apoptosis. Special attention will be given to oncogenes and tumor suppressor genes that are modulated by ionizing radiation and the tumor microenvironment. We will relate the biology of oncogenes and tumor suppressor genes to basic aspects of radiation biology that would be important in clinical practice. Finally, we will review recent studies on the prognostic significance of p53 mutations and apoptosis in tumor specimens. The main point of this lecture is to relate both researcher and clinician what are the therapeutic ramifications of oncogene and tumor suppressor gene mutations found in human neoptasia

[miR-143 inhibits cell proliferation through targeted regulating the expression of K-ras gene in HeLa cells].

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Qin, H X; Cui, H K; Pan, Y; Hu, R L; Zhu, L H; Wang, S J

2016-12-23

Objective: To explore the effect of microRNA miR-143 on the proliferation of cervical cancer HeLa cells through targeted regulating the expression of K-ras gene. Methods: The luciferase report carrier containing wild type 3'-UTR of K-ras gene (K-ras-wt) or mutated 3'-UTR of the K-ras (K-ras-mut) were co-transfected with iR-143 mimic into the HeLa cells respectively, and the targeting effect of miR-143 in the transfectants was verified by the dual luciferase report system. HeLa cells were also transfected with miR-143 mimic (miR-143 mimic group), mimic control (negative control group), and miR-143 mimic plus K-ras gene (miR-143 mimic+ K-ras group), respectively. The expression of miR-143 in the transfected HeLa cells was detected by real-time PCR (RT-PCR), and the expression of K-ras protein was detected by Western blot. The cell proliferation activity of each group was examined by MTT assay. In addition, human cervical cancer tissue samples ( n =5) and cervical intraepithelial neoplasia tissue samples ( n =5) were also examined for the expression of miR-143 and K-ras protein by RT-PCR and Western blot, respectively. Results: The luciferase report assay showed that co-transfection with miR-143 mimic decreased the luciferase activity of the K-ras-wt significantly, but did not inhibit the luciferase activity of the K-ras-mut. The expression of miR-143 in the HeLa cells transfected with miR-143 mimic was significantly higher than that in the HeLa cells transfected with the mimic control (3.31±0.45 vs 0.97±0.22, P cell proliferative activity of the miR-143 mimic group was significantly lower than that of the negative control group ( P cell proliferative activity of the miR-143 mimic+ K-ras group was also significantly lower than the control group ( P HeLa cells through targeted regulating the expression of K-ras gene. In human cervical cancer tissues of a small sample set, the expression of miR-143 is downregulated, and the expression of K-ras is upregulated.

HSF and Msn2/4p can exclusively or cooperatively activate the yeast HSP104 gene.

Science.gov (United States)

Grably, Melanie R; Stanhill, Ariel; Tell, Osnat; Engelberg, David

2002-04-01

In an effort to understand how an accurate level of stress-specific expression is obtained, we studied the promoter of the yeast HSP104 gene. Through 5' deletions, we defined a 334 bp fragment upstream of the first coding AUG as sufficient and essential for maximal basal activity and a 260 bp fragment as sufficient and essential for heat shock responsiveness. These sequences contain heat shock elements (HSEs) and stress response elements (STREs) that cooperate to achieve maximal inducible expression. However, in the absence of one set of factors (e.g. in msn2Deltamsn4Delta cells) proper induction is obtained exclusively through HSEs. We also show that HSP104 is constitutively derepressed in ras2Delta cells. This derepression is achieved exclusively through activation of STREs, with no role for HSEs. Strikingly, in ras2Deltamsn2Deltamsn4Delta cells the HSP104 promoter is also derepressed, but in this strain derepression is mediated through HSEs, showing the flexibility and adaptation of the promoter. Thus, appropriate transcription of HSP104 is usually obtained through cooperation between the Msn2/4/STRE and the HSF/ HSE systems, but each factor could activate the promoter alone, backing up the other. Transcription control of HSP104 is adaptive and robust, ensuring proper expression under extreme conditions and in various mutants.

Loss of p53 induces cell proliferation via Ras-independent activation of the Raf/Mek/Erk signaling pathway

Science.gov (United States)

Drosten, Matthias; Sum, Eleanor Y. M.; Lechuga, Carmen G.; Simón-Carrasco, Lucía; Jacob, Harrys K. C.; García-Medina, Raquel; Huang, Sidong; Beijersbergen, Roderick L.; Bernards, Rene; Barbacid, Mariano

2014-01-01

The Ras family of small GTPases constitutes a central node in the transmission of mitogenic stimuli to the cell cycle machinery. The ultimate receptor of these mitogenic signals is the retinoblastoma (Rb) family of pocket proteins, whose inactivation is a required step to license cell proliferation. However, little is known regarding the molecular events that connect Ras signaling with the cell cycle. Here, we provide genetic evidence to illustrate that the p53/p21 Cdk-interacting protein 1 (Cip1)/Rb axis is an essential component of the Ras signaling pathway. Indeed, knockdown of p53, p21Cip1, or Rb restores proliferative properties in cells arrested by ablation of the three Ras loci, H-, N- and K-Ras. Ras signaling selectively inactivates p53-mediated induction of p21Cip1 expression by inhibiting acetylation of specific lysine residues in the p53 DNA binding domain. Proliferation of cells lacking both Ras proteins and p53 can be prevented by reexpression of the human p53 ortholog, provided that it retains an active DNA binding domain and an intact lysine residue at position 164. These results unveil a previously unidentified role for p53 in preventing cell proliferation under unfavorable mitogenic conditions. Moreover, we provide evidence that cells lacking Ras and p53 proteins owe their proliferative properties to the unexpected retroactivation of the Raf/Mek/Erk cascade by a Ras-independent mechanism. PMID:25288756

Silencing Agrobacterium oncogenes in transgenic grapevine results in strain-specific crown gall resistance.

Science.gov (United States)

Galambos, A; Zok, A; Kuczmog, A; Oláh, R; Putnoky, P; Ream, W; Szegedi, E

2013-11-01

Grapevine rootstock transformed with an Agrobacterium oncogene-silencing transgene was resistant to certain Agrobacterium strains but sensitive to others. Thus, genetic diversity of Agrobacterium oncogenes may limit engineering crown gall resistance. Crown gall disease of grapevine induced by Agrobacterium vitis or Agrobacterium tumefaciens causes serious economic losses in viticulture. To establish crown gall-resistant lines, somatic proembryos of Vitis berlandieri × V. rupestris cv. 'Richter 110' rootstock were transformed with an oncogene-silencing transgene based on iaaM and ipt oncogene sequences from octopine-type, tumor-inducing (Ti) plasmid pTiA6. Twenty-one transgenic lines were selected, and their transgenic nature was confirmed by polymerase chain reaction (PCR). These lines were inoculated with two A. tumefaciens and three A. vitis strains. Eight lines showed resistance to octopine-type A. tumefaciens A348. Resistance correlated with the expression of the silencing genes. However, oncogene silencing was mostly sequence specific because these lines did not abolish tumorigenesis by A. vitis strains or nopaline-type A. tumefaciens C58.

H- ras deletion protects against angiotensin II-induced arterial hypertension and cardiac remodeling through protein kinase G-Iβ pathway activation.

Science.gov (United States)

Martín-Sánchez, Paloma; Luengo, Alicia; Griera, Mercedes; Orea, María Jesús; López-Olañeta, Marina; Chiloeches, Antonio; Lara-Pezzi, Enrique; de Frutos, Sergio; Rodríguez-Puyol, Manuel; Calleros, Laura; Rodríguez-Puyol, Diego

2018-02-01

Ras proteins regulate cell survival, growth, differentiation, blood pressure, and fibrosis in some organs. We have demonstrated that H- ras gene deletion produces mice hypotension via a soluble guanylate cyclase-protein kinase G (PKG)-dependent mechanism. In this study, we analyzed the consequences of H- ras deletion on cardiac remodeling induced by continuous angiotensin II (AngII) infusion and the molecular mechanisms implied. Left ventricular posterior wall thickness and mass and cardiomyocyte cross-sectional area were similar between AngII-treated H-Ras knockout (H -ras -/- ) and control wild-type (H -ras +/+ ) mice, as were extracellular matrix protein expression. Increased cardiac PKG-Iβ protein expression in H -ras -/- mice suggests the involvement of this protein in heart protection. Ex vivo experiments on cardiac explants could support this mechanism, as PKG blockade blunted protection against AngII-induced cardiac hypertrophy and fibrosis markers in H -ras -/- mice. Genetic modulation studies in cardiomyocytes and cardiac and embryonic fibroblasts revealed that the lack of H-Ras down-regulates the B-RAF/MEK/ERK pathway, which induces the glycogen synthase kinase-3β-dependent activation of the transcription factor, cAMP response element-binding protein, which is responsible for PKG-Iβ overexpression in H -ras -/- mouse embryonic fibroblasts. This study demonstrates that H- ras deletion protects against AngII-induced cardiac remodeling, possibly via a mechanism in which PKG-Iβ overexpression could play a partial role, and points to H-Ras and/or downstream proteins as potential therapeutic targets in cardiovascular disease.-Martín-Sánchez, P., Luengo, A., Griera, M., Orea, M. J., López-Olañeta, M., Chiloeches, A., Lara-Pezzi, E., de Frutos, S., Rodríguez-Puyol, M., Calleros, L., Rodríguez-Puyol, D. H- ras deletion protects against angiotensin II-induced arterial hypertension and cardiac remodeling through protein kinase G-Iβ pathway activation.

Retroviruses Hijack Chromatin Loops to Drive Oncogene Expression and Highlight the Chromatin Architecture around Proto-Oncogenic Loci

Science.gov (United States)

Pattison, Jillian M.; Wright, Jason B.; Cole, Michael D.

2015-01-01

The majority of the genome consists of intergenic and non-coding DNA sequences shown to play a major role in different gene regulatory networks. However, the specific potency of these distal elements as well as how these regions exert function across large genomic distances remains unclear. To address these unresolved issues, we closely examined the chromatin architecture around proto-oncogenic loci in the mouse and human genomes to demonstrate a functional role for chromatin looping in distal gene regulation. Using cell culture models, we show that tumorigenic retroviral integration sites within the mouse genome occur near existing large chromatin loops and that this chromatin architecture is maintained within the human genome as well. Significantly, as mutagenesis screens are not feasible in humans, we demonstrate a way to leverage existing screens in mice to identify disease relevant human enhancers and expose novel disease mechanisms. For instance, we characterize the epigenetic landscape upstream of the human Cyclin D1 locus to find multiple distal interactions that contribute to the complex cis-regulation of this cell cycle gene. Furthermore, we characterize a novel distal interaction upstream of the Cyclin D1 gene which provides mechanistic evidence for the abundant overexpression of Cyclin D1 occurring in multiple myeloma cells harboring a pathogenic translocation event. Through use of mapped retroviral integrations and translocation breakpoints, our studies highlight the importance of chromatin looping in oncogene expression, elucidate the epigenetic mechanisms crucial for distal cis-regulation, and in one particular instance, explain how a translocation event drives tumorigenesis through upregulation of a proto-oncogene. PMID:25799187

bcr-abl oncogene activation in Philadelphia chromosome-positive acute lymphoblastic leukemia

NARCIS (Netherlands)

Hermans, A.; Gow, J.; Selleri, L.; von Lindern, M.; Hagemeijer, A.; Wiedemann, L. M.; Grosveld, G.

1988-01-01

Tumor-specific alterations in oncogenes are thought to play a central role in the development of cancer. An example is the consistent fusion of the bcr gene to the c-abl oncogene on the Ph chromosome in CML. The Ph chromosome can also be observed in ALL. About 50% of Ph+ ALL cases, in contrast to

The regulation of ras-raf signaling pathway on G1 phase of the irradiated cells

International Nuclear Information System (INIS)

Guo Dehuang; Dong Bo; Liu Nongle; Wen Gengyun; Luo Qingliang; Mao Bingzhi

2000-01-01

Objective: To investigate the way of ras-raf signaling pathway which regulate the G 1 phase in irradiated KG-1 cells. Methods: Blocked the GM-CSF signaling pathway by transfected DN-ras and then momentary transfected cyclin D1 into irradiated KG-1 cells, the effects of cyclin D1 on G 1 phase was examined. Results: The irradiated KG-1 cells transfected DN-ras can't recover form G 1 phase arrest even though the GM-CSF was given,momentary transfected cyclin D1 promote the irradiated KG-1 cells from G 1 arrest. Conclusion: Activation of ras-raf signaling pathway regulate the cell cycle of the irradiated KG-1 cells through promotion the expression of the cyclin D1

Alphavirus production is inhibited in neurofibromin 1-deficient cells through activated RAS signalling

International Nuclear Information System (INIS)

Kolokoltsova, Olga A.; Domina, Aaron M.; Kolokoltsov, Andrey A.; Davey, Robert A.; Weaver, Scott C.; Watowich, Stanley J.

2008-01-01

Virus-host interactions essential for alphavirus pathogenesis are poorly understood. To address this shortcoming, we coupled retrovirus insertional mutagenesis and a cell survival selection strategy to generate clonal cell lines broadly resistant to Sindbis virus (SINV) and other alphaviruses. Resistant cells had significantly impaired SINV production relative to wild-type (WT) cells, although virus binding and fusion events were similar in both sets of cells. Analysis of the retroviral integration sites identified the neurofibromin 1 (NF1) gene as disrupted in alphavirus-resistant cell lines. Subsequent analysis indicated that expression of NF1 was significantly reduced in alphavirus-resistant cells. Importantly, independent down-regulation of NF1 expression in WT HEK 293 cells decreased virus production and increased cell viability during SINV infection, relative to infected WT cells. Additionally, we observed hyperactive RAS signalling in the resistant HEK 293 cells, which was anticipated because NF1 is a negative regulator of RAS. Expression of constitutively active RAS (HRAS-G12V) in a WT HEK 293 cell line resulted in a marked delay in virus production, compared with infected cells transfected with parental plasmid or dominant-negative RAS (HRAS-S17N). This work highlights novel host cell determinants required for alphavirus pathogenesis and suggests that RAS signalling may play an important role in neuronal susceptibility to SINV infection

Ras Signaling Regulates Stem Cells and Amelogenesis in the Mouse Incisor.

Science.gov (United States)

Zheng, X; Goodwin, A F; Tian, H; Jheon, A H; Klein, O D

2017-11-01

The role of Ras signaling during tooth development is poorly understood. Ras proteins-which are activated by many upstream pathways, including receptor tyrosine kinase cascades-signal through multiple effectors, such as the mitogen-activated protein kinase (MAPK) and PI3K pathways. Here, we utilized the mouse incisor as a model to study how the MAPK and PI3K pathways regulate dental epithelial stem cells and amelogenesis. The rodent incisor-which grows continuously throughout the life of the animal due to the presence of epithelial and mesenchymal stem cells-provides a model for the study of ectodermal organ renewal and regeneration. Utilizing models of Ras dysregulation as well as inhibitors of the MAPK and PI3K pathways, we found that MAPK and PI3K regulate dental epithelial stem cell activity, transit-amplifying cell proliferation, and enamel formation in the mouse incisor.

FGFR3, PIK3CA and RAS mutations in benign lichenoid keratosis.

Science.gov (United States)

Groesser, L; Herschberger, E; Landthaler, M; Hafner, C

2012-04-01

Benign lichenoid keratoses (BLKs) are solitary skin lesions which have been proposed to represent a regressive form of pre-existent epidermal tumours such as solar lentigo or seborrhoeic keratosis. However, the genetic basis of BLK is unknown. FGFR3, PIK3CA and RAS mutations have been shown to be involved in the pathogenesis of seborrhoeic keratosis and solar lentigo. We thus investigated whether these mutations are also present in BLK. After manual microdissection and DNA isolation, 52 BLKs were screened for FGFR3, PIK3CA and RAS hotspot mutations using SNaPshot(®) multiplex assays. We identified 6/52 (12%) FGFR3 mutations, 10/52 (19%) PIK3CA mutations, 6/52 (12%) HRAS mutations and 2/52 (4%) KRAS mutations. FGFR3 and RAS mutations were mutually exclusive. One BLK showed a simultaneous PIK3CA and HRAS mutation. In nine BLKs with a mutation, nonlesional control tissue from the epidermal margin and the dermal lymphocytic infiltrate were wild-type, indicating that these mutations are somatic. To demonstrate that these findings are specific, 10 samples of lichen planus were analysed without evidence for FGFR3, PIK3CA or RAS mutations. Our results indicate that FGFR3, PIK3CA and RAS mutations are present in approximately 50% of BLKs. These findings support the concept on the molecular genetic level that at least a proportion of BLKs represents regressive variants resulting from former benign epidermal tumours such as seborrhoeic keratosis and solar lentigo. © 2011 The Authors. BJD © 2011 British Association of Dermatologists 2011.

Early bichemical markers of effects: Enzyme induction, oncogene activation and markers of oxidative damage

DEFF Research Database (Denmark)

Poulsen, Henrik E.; Loft, Steffen

1995-01-01

Early bichemical marker, enzyme induction, oncogene activation, oxidative damage, low-density lipoprotein......Early bichemical marker, enzyme induction, oncogene activation, oxidative damage, low-density lipoprotein...

Thermodynamics between RAP/RAS and virgin aggregates during asphalt concrete production : a literature review.

Science.gov (United States)

2015-09-01

In hot-mix asphalt (HMA) plants, virgin aggregates are heated and dried separately before being mixed with : RAP/RAS and virgin asphalt binder. RAP/RAS materials are not heated or dried directly by a burner to avoid : burning of aged binder coating o...

Active ras triggers death in glioblastoma cells through hyperstimulation of macropinocytosis.

Science.gov (United States)

Overmeyer, Jean H; Kaul, Aparna; Johnson, Erin E; Maltese, William A

2008-06-01

Expression of activated Ras in glioblastoma cells induces accumulation of large phase-lucent cytoplasmic vacuoles, followed by cell death. This was previously described as autophagic cell death. However, unlike autophagosomes, the Ras-induced vacuoles are not bounded by a double membrane and do not sequester organelles or cytoplasm. Moreover, they are not acidic and do not contain the autophagosomal membrane protein LC3-II. Here we show that the vacuoles are enlarged macropinosomes. They rapidly incorporate extracellular fluid-phase tracers but do not sequester transferrin or the endosomal protein EEA1. Ultimately, the cells expressing activated Ras detach from the substratum and rupture, coincident with the displacement of cytoplasm with huge macropinosome-derived vacuoles. These changes are accompanied by caspase activation, but the broad-spectrum caspase inhibitor carbobenzoxy-Val-Ala-Asp-fluoromethylketone does not prevent cell death. Moreover, the majority of degenerating cells do not exhibit chromatin condensation typical of apoptosis. These observations provide evidence for a necrosis-like form of cell death initiated by dysregulation of macropinocytosis, which we have dubbed "methuosis." An activated form of the Rac1 GTPase induces a similar form of cell death, suggesting that Ras acts through Rac-dependent signaling pathways to hyperstimulate macropinocytosis in glioblastoma. Further study of these signaling pathways may lead to the identification of other chemical and physiologic triggers for this unusual form of cell death.

Rasfonin, a novel 2-pyrone derivative, induces ras-mutated Panc-1 pancreatic tumor cell death in nude mice.

Science.gov (United States)

Xiao, Z; Li, L; Li, Y; Zhou, W; Cheng, J; Liu, F; Zheng, P; Zhang, Y; Che, Y

2014-05-22

Rasfonin is a novel 2-pyrone derivative reported to induce apoptosis in ras-dependent cells. In this study, its effects on ras-mutated pancreatic cancer cells were investigated in vitro and in vivo. Two human pancreatic cancer cell lines Panc-1 (mutated K-ras) and BxPC-3 (wild-type K-ras) were selected to test the effects of rasfonin on cell proliferation, clone formation, migration and invasion in vitro. Immunoblotting was used to detect the expressions of EGFR-Ras-Raf-MEK-ERK signaling pathway proteins. Ras activity was measured using a pull-down ELISA kit and guanine exchange factor (GEF)/GTPase-activating proteins (GAP) activity was measured by [(3)H]-GDP radiometric ligand binding. For an in vivo study, CD1 nude mice bearing Panc-1 cells were treated with rasfonin or Salirasib (FTS). We found that rasfonin suppressed proliferation more strongly in Panc-1 cells (IC50=5.5 μM) than BxPC-3 cells (IC50=10 μM) in vitro. Clone formation, migration and invasion by Panc-1 cells were also reduced by rasfonin. Rasfonin had little effect on the farnesylation of Ras, but it strongly downregulated Ras activity and consequently phosphorylation of c-Raf/MEK/ERK. Further experiments indicated that rasfonin reduced Son of sevenless (Sos1) expression but did not alter GEF and GAP activities. The in vivo experiments also revealed that rasfonin (30 mg/kg) delayed the growth of xenograft tumors originating from Panc-1 cells. Tumor weight was ultimately decreased after 20 days of treatment of rasfonin. Rasfonin is a robust inhibitor of pancreatic cancers with the K-ras mutation. The reduction of Sos1 expression and the consequently depressed Ras-MAPK activity could be important in its anticancer activity.

Absence of mutations in the coding sequence of the potential tumor suppressor 3pK in metastatic melanoma

Directory of Open Access Journals (Sweden)

Houben Roland

2005-12-01

Full Text Available Abstract Background Activation of Ras or Raf contributes to tumorigenesis of melanoma. However, constitutive Raf activation is also a characteristic of the majority of benign melanocytic nevi and high intensity signaling of either Ras or Raf was found to induce growth inhibition and senescence rather than transformation. Since the chromosome 3p kinase (3pK is a target of the Ras/Raf/Mek/Erk signaling pathway which antagonizes the function of the oncogene and anti-differentiation factor Bmi-1, 3pK may function as a tumor suppressor in tumors with constitutive Ras/Raf activation. Consequently, we tested whether inactivating 3pK mutations are present in melanoma. Methods 30 metastatic melanoma samples, which were positive for activating mutations of either BRaf or NRas, were analyzed for possible mutations in the 3pk gene. The 10 coding exons and their flanking intron sequences were amplified by PCR and direct sequencing of the PCR products was performed. Results This analysis revealed that besides the presence of some single nucleotide polymorphisms in the 3pk gene, we could not detect any possible loss of function mutation in any of these 30 metastatic melanoma samples selected for the presence of activating mutations within the Ras/Raf/Mek/Erk signaling pathway. Conclusion Hence, in melanoma with constitutively active Ras/Raf inactivating mutations within the 3pk gene do not contribute to the oncogenic phenotype of this highly malignant tumor.

Ras-dva is a novel Pit-1- and glucocorticoid-regulated gene in the embryonic anterior pituitary gland.

Science.gov (United States)

Ellestad, Laura E; Porter, Tom E

2013-01-01

Glucocorticoids play a role in functional differentiation of pituitary somatotrophs and lactotrophs during embryogenesis. Ras-dva was identified as a gene regulated by anterior neural fold protein-1/homeobox expressed in embryonic stem cells-1, a transcription factor known to be critical in pituitary development, and has an expression profile in the chicken embryonic pituitary gland that is consistent with in vivo regulation by glucocorticoids. The objective of this study was to characterize expression and regulation of ras-dva mRNA in the developing chicken anterior pituitary. Pituitary ras-dva mRNA levels increased during embryogenesis to a maximum on embryonic day (e) 18 and then decreased and remained low or undetectable after hatch. Ras-dva expression was highly enriched in the pituitary gland on e18 relative to other tissues examined. Glucocorticoid treatment of pituitary cells from mid- and late-stage embryos rapidly increased ras-dva mRNA, suggesting it may be a direct transcriptional target of glucocorticoids. A reporter construct driven by 4 kb of the chicken ras-dva 5'-flanking region, containing six putative pituitary-specific transcription factor-1 (Pit-1) binding sites and two potential glucocorticoid receptor (GR) binding sites, was highly activated in embryonic pituitary cells and up-regulated by corticosterone. Mutagenesis of the most proximal Pit-1 site decreased promoter activity in chicken e11 pituitary cells, indicating regulation of ras-dva by Pit-1. However, mutating putative GR binding sites did not substantially reduce induction of ras-dva promoter activity by corticosterone, suggesting additional DNA elements within the 5'-flanking region are responsible for glucocorticoid regulation. We have identified ras-dva as a glucocorticoid-regulated gene that is likely expressed in cells of the Pit-1 lineage within the developing anterior pituitary gland.

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...

Dysregulated ΔNp63α inhibits expression of Ink4a/arf, blocks senescence, and promotes malignant conversion of keratinocytes.

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Linan Ha

Full Text Available p63 is critical for squamous epithelial development, and elevated levels of the ΔNp63α isoform are seen in squamous cell cancers of various organ sites. However, significant controversy exists regarding the role of p63 isoforms as oncoproteins or tumor suppressors. Here, lentiviruses were developed to drive long-term overexpression of ΔNp63α in primary keratinocytes. Elevated levels of ΔNp63α in vitro promote long-term survival and block both replicative and oncogene-induced senescence in primary keratinocytes, as evidenced by the expression of SA-β-gal and the presence of nuclear foci of heterochromatin protein 1γ. The contribution of ΔNp63α to cancer development was assessed using an in vivo grafting model of experimental skin tumorigenesis that allows distinction between benign and malignant tumors. Grafted lenti-ΔNp63α keratinocytes do not form tumors, whereas lenti-GFP/v-ras(Ha keratinocytes develop well-differentiated papillomas. Lenti-ΔNp63α/v-ras(Ha keratinocytes form undifferentiated carcinomas. The average volume of lenti-ΔNp63α/v-ras(Ha tumors was significantly higher than those in the lenti-GFP/v-ras(Ha group, consistent with increased BrdU incorporation detected by immunohistochemistry. The block in oncogene-induced senescence corresponds to sustained levels of E2F1 and phosphorylated AKT, and is associated with loss of induction of p16(ink4a/p19(arf. The relevance of p16(ink4a/p19(arf loss was demonstrated in grafting studies of p19(arf-null keratinocytes, which develop malignant carcinomas in the presence of v-ras(Ha similar to those arising in wildtype keratinocytes that express lenti-ΔNp63α and v-ras(Ha. Our findings establish that ΔNp63α has oncogenic activity and its overexpression in human squamous cell carcinomas contributes to the malignant phenotype, and implicate its ability to regulate p16(ink4a/p19(arf in the process.

Clasificación de presas y evaluación del riesgo con programa HEC-RAS

OpenAIRE

Marín Rubís, Àlex

2006-01-01

Desde el año 2003 el modelo HEC-RAS, desarrollado por el Hydrologic Engineering Center (HEC) del United States Army Corps of Engineers, incorpora entre sus capacidades un módulo que permite la simulación de rotura de presas. El objeto de este trabajo ha sido asegurar la capacidad de HEC-RAS para llevar a cabo este tipo de simulaciones y crear una guía para su uso en entorno GIS (Arcview) mediante la aplicación HEC-GeoRAS.

The Ras GTPase-activating protein Rasal3 supports survival of naive T cells.

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Ryunosuke Muro

Full Text Available The Ras-mitogen-activated protein kinase (MAPK pathway is crucial for T cell receptor (TCR signaling in the development and function of T cells. The significance of various modulators of the Ras-MAPK pathway in T cells, however, remains to be fully understood. Ras-activating protein-like 3 (Rasal3 is an uncharacterized member of the SynGAP family that contains a conserved Ras GTPase-activating protein (GAP domain, and is predominantly expressed in the T cell lineage. In the current study, we investigated the function and physiological roles of Rasal3. Our results showed that Rasal3 possesses RasGAP activity, but not Rap1GAP activity, and represses TCR-stimulated ERK phosphorylation in a T cell line. In systemic Rasal3-deficient mice, T cell development in the thymus including positive selection, negative selection, and β-selection was unaffected. However, the number of naive, but not effector memory CD4 and CD8 T cell in the periphery was significantly reduced in Rasal3-deficient mice, and associated with a marked increase in apoptosis of these cells. Indeed, survival of Rasal3 deficient naive CD4 T cells in vivo by adoptive transfer was significantly impaired, whereas IL-7-dependent survival of naive CD4 T cells in vitro was unaltered. Collectively, Rasal3 is required for in vivo survival of peripheral naive T cells, contributing to the maintenance of optimal T cell numbers.

Application of HEC-RAS water quality model to estimate contaminant spreading in small stream

Energy Technology Data Exchange (ETDEWEB)

Halaj, Peter; Bárek, Viliam; Halajová, Anna Báreková; Halajová, Denisa [Slovak University of Agriculture in Nitra, Nitra (Slovakia)

2013-07-01

The paper presents study of some aspects of HEC-RAS water quality model connected to simulation of contaminant transport in small stream. Authors mainly focused on one of the key tasks in process of pollutant transport modelling in streams - determination of the dispersion characteristics represented by longitudinal dispersion coefficient D. Different theoretical and empirical formulas have been proposed for D value determination and they have revealed that the coefficient is variable parameter that depends on hydraulic and morphometric characteristics of the stream reaches. Authors compare the results of several methods of coefficient D assessment, assuming experimental data obtained by tracer studies and compare them with results optimized by HEC-RAS water quality model. The analyses of tracer study and computation outputs allow us to outline the important aspects of longitudinal dispersion coefficient set up in process of the HEC-RAS model use. Key words: longitudinal dispersion coefficient, HEC-RAS, water quality modeling.

Oncogenic HPV among HIV infected female population in West Bengal, India

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Sengupta Sharmila

2011-03-01

Full Text Available Abstract Background Prevalence of both cervical cancer and Human Immunodeficiency Virus (HIV infection are very high in India. Natural history of Human Papilloma Virus (HPV infection is known to be altered in HIV positive women and there is an increased possibility of persistence of HPV infections in this population. Therefore, this study was conducted to understand the epidemiology and circulating genotypes of oncogenic HPV among HIV positive and negative female population in West Bengal, India. Methods In this hospital-based cross-sectional study, 93 known HIV positive females attending a pre-ART registration clinic and 1106 HIV negative females attending a Reproductive and Child Health Care Clinic were subjected to study. Cervical cell samples collected from the study population were tested for the presence of HPV 16, 18 using specific primers. Roche PCR assay was used to detect other specific HPV genotypes in the cervical cells specimens of HIV positive cases only. Results Prevalence of HPV 16, 18 among HIV positive females (32.2%; n = 30 was higher than HIV negative females (9.1%; n = 101. About 53% (23/43 of cases with oncogenic HPV were infected with genotypes other than 16, 18 either as single/multiple infections. HPV 18 and HPV 16 were the predominant genotypes among HIV positive and HIV negative subjects respectively. Oncogenic HPV was not found to be associated with age and duration of sexual exposure. But the presence of HIV was found to a statistically significant predictor oncogenic HPV. Conclusion The currently available HPV vaccines offer protection only against HPV 16 and 18 and some cross- protection to few associated genotypes. These vaccines are therefore less likely to offer protection against cervical cancer in HIV positive women a high percentage of who were infected with non-16 and non-18 oncogenic HPV genotypes. Additionally, there is a lack of sufficient evidence of immunogenicity in HIV infected individuals. Therefore

Monitoring Ras Interactions with the Nucleotide Exchange Factor Son of Sevenless (Sos) Using Site-specific NMR Reporter Signals and Intrinsic Fluorescence*

Science.gov (United States)

Vo, Uybach; Vajpai, Navratna; Flavell, Liz; Bobby, Romel; Breeze, Alexander L.; Embrey, Kevin J.; Golovanov, Alexander P.

2016-01-01

The activity of Ras is controlled by the interconversion between GTP- and GDP-bound forms partly regulated by the binding of the guanine nucleotide exchange factor Son of Sevenless (Sos). The details of Sos binding, leading to nucleotide exchange and subsequent dissociation of the complex, are not completely understood. Here, we used uniformly 15N-labeled Ras as well as [13C]methyl-Met,Ile-labeled Sos for observing site-specific details of Ras-Sos interactions in solution. Binding of various forms of Ras (loaded with GDP and mimics of GTP or nucleotide-free) at the allosteric and catalytic sites of Sos was comprehensively characterized by monitoring signal perturbations in the NMR spectra. The overall affinity of binding between these protein variants as well as their selected functional mutants was also investigated using intrinsic fluorescence. The data support a positive feedback activation of Sos by Ras·GTP with Ras·GTP binding as a substrate for the catalytic site of activated Sos more weakly than Ras·GDP, suggesting that Sos should actively promote unidirectional GDP → GTP exchange on Ras in preference of passive homonucleotide exchange. Ras·GDP weakly binds to the catalytic but not to the allosteric site of Sos. This confirms that Ras·GDP cannot properly activate Sos at the allosteric site. The novel site-specific assay described may be useful for design of drugs aimed at perturbing Ras-Sos interactions. PMID:26565026

miR-11 regulates pupal size of Drosophila melanogaster via directly targeting Ras85D.

Science.gov (United States)

Li, Yao; Li, Shengjie; Jin, Ping; Chen, Liming; Ma, Fei

2017-01-01

MicroRNAs play diverse roles in various physiological processes during Drosophila development. In the present study, we reported that miR-11 regulates pupal size during Drosophila metamorphosis via targeting Ras85D with the following evidences: pupal size was increased in the miR-11 deletion mutant; restoration of miR-11 in the miR-11 deletion mutant rescued the increased pupal size phenotype observed in the miR-11 deletion mutant; ectopic expression of miR-11 in brain insulin-producing cells (IPCs) and whole body shows consistent alteration of pupal size; Dilps and Ras85D expressions were negatively regulated by miR-11 in vivo; miR-11 targets Ras85D through directly binding to Ras85D 3'-untranslated region in vitro; removal of one copy of Ras85D in the miR-11 deletion mutant rescued the increased pupal size phenotype observed in the miR-11 deletion mutant. Thus, our current work provides a novel mechanism of pupal size determination by microRNAs during Drosophila melanogaster metamorphosis. Copyright © 2017 the American Physiological Society.

Nucleotide sequence of a human cDNA encoding a ras-related protein (rap1B)

Energy Technology Data Exchange (ETDEWEB)

Pizon, V; Lerosey, I; Chardin, P; Tavitian, A [INSERM, Paris (France)

1988-08-11

The authors have previously characterized two human ras-related genes rap1 and rap2. Using the rap1 clone as probe they isolated and sequenced a new rap cDNA encoding the 184aa rap1B protein. The rap1B protein is 95% identical to rap1 and shares several properties with the ras protein suggesting that it could bind GTP/GDP and have a membrane location. As for rap1, the structural characteristics of rap1B suggest that the rap and ras proteins might interact on the same effector.

Using 18F FDG PET/CT to Detect an occult Mesenchymal Tumor Causing Oncogenic Osteomalacia

International Nuclear Information System (INIS)

Seo, Hyo Jung; Choi, Yun Jung; Kim, Hyun Jeong; Jeong, Yong Hyu; Cho, Arthur; Lee, Jae Hoon; Yun, Mijin; Lee, Jong Doo; Kang, Won Jun

2011-01-01

Oncogenic osteomalacia is a rare paraneoplastic syndrome characterized by renal phosphate excretion, hypophosphatemia, and osteomalacia. This syndrome is often caused by tumors of mesenchymal origin. Patients with oncogenic osteomalacia have abnormal bone mineralization, resulting in a high frequency of fractures. Tumor resection is the treatment of choice, as it will often correct the metabolic imbalance. Although oncogenic osteomalacia is a potentially curable disease, diagnosis is difficult and often delayed because of the small size and sporadic location of the tumor. Bone scintigraphy and radiography best characterize osteoma lacia; magnetic resonance imaging findings are nonspecific. Here, we report a case of oncogenic osteomalacia secondary to a phosphaturic mesenchymal tumor that was successfully detected by 18F fluorodeoxyglucose positron emission tomography/computed tomography ( 18F FDG PET/CT). This case illustrates the advantages of 18F FDG PET/CT in detecting the occult mesenchymal tumor that causes oncogenic osteomalacia.

Exosomes facilitate therapeutic targeting of oncogenic KRAS in pancreatic cancer.

Science.gov (United States)

Kamerkar, Sushrut; LeBleu, Valerie S; Sugimoto, Hikaru; Yang, Sujuan; Ruivo, Carolina F; Melo, Sonia A; Lee, J Jack; Kalluri, Raghu

2017-06-22

The mutant form of the GTPase KRAS is a key driver of pancreatic cancer but remains a challenging therapeutic target. Exosomes are extracellular vesicles generated by all cells, and are naturally present in the blood. Here we show that enhanced retention of exosomes, compared to liposomes, in the circulation of mice is likely due to CD47-mediated protection of exosomes from phagocytosis by monocytes and macrophages. Exosomes derived from normal fibroblast-like mesenchymal cells were engineered to carry short interfering RNA or short hairpin RNA specific to oncogenic Kras G12D , a common mutation in pancreatic cancer. Compared to liposomes, the engineered exosomes (known as iExosomes) target oncogenic KRAS with an enhanced efficacy that is dependent on CD47, and is facilitated by macropinocytosis. Treatment with iExosomes suppressed cancer in multiple mouse models of pancreatic cancer and significantly increased overall survival. Our results demonstrate an approach for direct and specific targeting of oncogenic KRAS in tumours using iExosomes.

Combating oncogene activation associated with retrovirus-mediated gene therapy of X-linked severe combined immunodeficiency

Directory of Open Access Journals (Sweden)

B.E. Strauss

2007-05-01

Full Text Available A successful gene therapy clinical trial that also encountered serious adverse effects has sparked extensive study and debate about the future directions for retrovirus-mediated interventions. Treatment of X-linked severe combined immunodeficiency with an oncoretrovirus harboring a normal copy of the gc gene was applied in two clinical trials, essentially curing 13 of 16 infants, restoring a normal immune system without the need for additional immune-related therapies. Approximately 3 years after their gene therapy, tragically, 3 of these children, all from the same trial, developed leukemia as a result of this experimental treatment. The current understanding of the mechanism behind this leukemogenesis involves three critical and cooperating factors, i.e., viral integration, oncogene activation, and the function of the therapeutic gene. In this review, we will explore the causes of this unwanted event and some of the possibilities for reducing the risk of its reoccurrence.

K-ras mutations in sinonasal cancers in relation to wood dust exposure

International Nuclear Information System (INIS)

Bornholdt, Jette; Vogel, Ulla; Husgafvel-Pursiainen, Kirsti; Wallin, Håkan; Hansen, Johnni; Steiniche, Torben; Dictor, Michael; Antonsen, Annemarie; Wolff, Henrik; Schlünssen, Vivi; Holmila, Reetta; Luce, Danièle

2008-01-01

Cancer in the sinonasal tract is rare, but persons who have been occupationally exposed to wood dust have a substantially increased risk. It has been estimated that approximately 3.6 million workers are exposed to inhalable wood dust in EU. In previous small studies of this cancer, ras mutations were suggested to be related to wood dust exposure, but these studies were too limited to detect statistically significant associations. We examined 174 cases of sinonasal cancer diagnosed in Denmark in the period from 1991 to 2001. To ensure uniformity, all histological diagnoses were carefully reviewed pathologically before inclusion. Paraffin embedded tumour samples from 58 adenocarcinomas, 109 squamous cell carcinomas and 7 other carcinomas were analysed for K-ras codon 12, 13 and 61 point mutations by restriction fragment length polymorphisms and direct sequencing. Information on occupational exposure to wood dust and to potential confounders was obtained from telephone interviews and from registry data. Among the patients in this study, exposure to wood dust was associated with a 21-fold increased risk of having an adenocarcinoma than a squamous cell carcinoma compared to unexposed [OR = 21.0, CI = 8.0–55.0]. K-ras was mutated in 13% of the adenocarcinomas (seven patients) and in 1% of squamous cell carcinomas (one patient). Of these eight mutations, five mutations were located in the codon 12. The exact sequence change of remaining three could not be identified unambiguously. Among the five identified mutations, the G→A transition was the most common, and it was present in tumour tissue from two wood dust exposed adenocarcinoma patients and one patient with unknown exposure. Previously published studies of sinonasal cancer also identify the GGT → GAT transition as the most common and often related to wood dust exposure. Patients exposed to wood dust seemed more likely to develop adenocarcinoma compared to squamous cell carcinomas. K-ras mutations were detected

Monitoring Ras Interactions with the Nucleotide Exchange Factor Son of Sevenless (Sos) Using Site-specific NMR Reporter Signals and Intrinsic Fluorescence.

Science.gov (United States)

Vo, Uybach; Vajpai, Navratna; Flavell, Liz; Bobby, Romel; Breeze, Alexander L; Embrey, Kevin J; Golovanov, Alexander P

2016-01-22

The activity of Ras is controlled by the interconversion between GTP- and GDP-bound forms partly regulated by the binding of the guanine nucleotide exchange factor Son of Sevenless (Sos). The details of Sos binding, leading to nucleotide exchange and subsequent dissociation of the complex, are not completely understood. Here, we used uniformly (15)N-labeled Ras as well as [(13)C]methyl-Met,Ile-labeled Sos for observing site-specific details of Ras-Sos interactions in solution. Binding of various forms of Ras (loaded with GDP and mimics of GTP or nucleotide-free) at the allosteric and catalytic sites of Sos was comprehensively characterized by monitoring signal perturbations in the NMR spectra. The overall affinity of binding between these protein variants as well as their selected functional mutants was also investigated using intrinsic fluorescence. The data support a positive feedback activation of Sos by Ras·GTP with Ras·GTP binding as a substrate for the catalytic site of activated Sos more weakly than Ras·GDP, suggesting that Sos should actively promote unidirectional GDP → GTP exchange on Ras in preference of passive homonucleotide exchange. Ras·GDP weakly binds to the catalytic but not to the allosteric site of Sos. This confirms that Ras·GDP cannot properly activate Sos at the allosteric site. The novel site-specific assay described may be useful for design of drugs aimed at perturbing Ras-Sos interactions. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Nitric oxide induces thioredoxin-1 nuclear translocation: Possible association with the p21Ras survival pathway

International Nuclear Information System (INIS)

Arai, Roberto J.; Masutani, H.; Yodoi, J.; Debbas, V.; Laurindo, Francisco R.; Stern, A.; Monteiro, Hugo P.

2006-01-01

One of the major redox-regulating molecules with thiol reducing activity is thioredoxin-1 (TRX-1). TRX-1 is a multifunctional protein that exists in the extracellular millieu, cytoplasm, and nucleus, and has a distinct role in each environment. It is well known that TRX-1 promptly migrates to the nuclear compartment in cells exposed to oxidants. However, the intracellular location of TRX-1 in cells exposed to nitrosothiols has not been investigated. Here, we demonstrated that the exposure of HeLa cells to increasing concentrations of the nitrosothiol S-nitroso-N-acetylpenicillamine (SNAP) promoted TRX-1 nuclear accumulation. The SNAP-induced TRX-1 translocation to the nucleus was inhibited by FPTIII, a selective inhibitor of p21Ras. Furthermore, TRX-1 migration was attenuated in cells stably transfected with NO insensitive p21Ras (p21 RasC118S ). Downstream to p21Ras, the MAP Kinases ERK1/2 were activated by SNAP under conditions that promote TRX-1 nuclear translocation. Inhibition of MEK prevented SNAP-stimulated ERK1/2 activation and TRX-1 nuclear migration. In addition, cells treated with p21Ras or MEK inhibitor showed increased susceptibility to cell death induced by SNAP. In conclusion, our observations suggest that the nuclear translocation of TRX-1 is induced by SNAP involving p21Ras survival pathway

Structural insight into the rearrangement of the switch I region in GTP-bound G12A K-Ras

Energy Technology Data Exchange (ETDEWEB)

Xu, Shenyuan; Long, Brian N.; Boris, Gabriel H.; Chen, Anqi; Ni, Shuisong; Kennedy, Michael A.

2017-11-10

K-Ras, a molecular switch that regulates cell growth, apoptosis and metabolism, is activated when it undergoes a conformation change upon binding GTP and is deactivated following the hydrolysis of GTP to GDP. Hydrolysis of GTP in water is accelerated by coordination to K-Ras, where GTP adopts a high-energy conformation approaching the transition state. The G12A mutation reduces intrinsic K-Ras GTP hydrolysis by an unexplained mechanism. Here, crystal structures of G12A K-Ras in complex with GDP, GTP, GTPγS and GppNHp, and of Q61A K-Ras in complex with GDP, are reported. In the G12A K-Ras–GTP complex, the switch I region undergoes a significant reorganization such that the Tyr32 side chain points towards the GTP-binding pocket and forms a hydrogen bond to the GTP γ-phosphate, effectively stabilizing GTP in its precatalytic state, increasing the activation energy required to reach the transition state and contributing to the reduced intrinsic GTPase activity of G12A K-Ras mutants.

Intracortical osteoblastic osteosarcoma with oncogenic rickets

International Nuclear Information System (INIS)

Hasegawa, T.; Hirohashi, Setsuo; Shimoda, Tadakazu; Yokoyama, Ryohei; Beppu, Yasuo; Maeda, Shotaro

1999-01-01

Intracortical osteosarcoma is the rarest variant of osteosarcoma, occurring within, and usually confined to, the cortical bone. Oncogenic osteomalacia, or rickets, is an unusual clinicopathologic entity in which vitamin D-resistant osteomalacia, or rickets, occurs in association with some tumors of soft tissue or bone. We present a case of oncogenic rickets associated with intracortical osteosarcoma of the tibia in a 9-year-old boy, whose roentgenographic abnormalities of rickets disappeared and pertinent laboratory data except for serum alkaline phosphatase became normal after surgical resection of the tumor. Histologically, the tumor was an osteosarcoma with a prominent osteoblastic pattern. An unusual microscopic feature was the presence of matrix mineralization showing rounded calcified structures (calcified spherules). Benign osteoblastic tumors, such as osteoid osteoma and osteoblastoma, must be considered in the differential diagnosis because of the relatively low cellular atypia and mitotic activity of this tumor. The infiltrating pattern with destruction or engulfment of normal bone is a major clue to the correct diagnosis of intracortical osteosarcoma. The co-existing radiographic changes of rickets were due to the intracortical osteosarcoma. (orig.)

Genetic and Molecular Analysis of Suppressors of Ras Mutations

National Research Council Canada - National Science Library

Sieburth, Derek

1998-01-01

.... elegans vulval development. We describe the identification and characterization of a novel gene, sur-8, that functions to regulate a receptor tyrosine kinase-Ras-MAP kinase-mediated signal transduction pathway during C...

Parallel imaging of Drosophila embryos for quantitative analysis of genetic perturbations of the Ras pathway

Directory of Open Access Journals (Sweden)

Yogesh Goyal

2017-07-01

Full Text Available The Ras pathway patterns the poles of the Drosophila embryo by downregulating the levels and activity of a DNA-binding transcriptional repressor Capicua (Cic. We demonstrate that the spatiotemporal pattern of Cic during this signaling event can be harnessed for functional studies of mutations in the Ras pathway in human diseases. Our approach relies on a new microfluidic device that enables parallel imaging of Cic dynamics in dozens of live embryos. We found that although the pattern of Cic in early embryos is complex, it can be accurately approximated by a product of one spatial profile and one time-dependent amplitude. Analysis of these functions of space and time alone reveals the differential effects of mutations within the Ras pathway. Given the highly conserved nature of Ras-dependent control of Cic, our approach provides new opportunities for functional analysis of multiple sequence variants from developmental abnormalities and cancers.