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Sample records for imaging oncogene expression

  1. Oncogenes

    SciTech Connect

    Compans, R.W.; Cooper, M.; Koprowski, H.; McConell, I.; Melchers, F.; Nussenzweig, V.; Oldstone, M.; Olsnes, S.; Saedler, H.; Vogt, P.K.

    1989-01-01

    This book covers the following topics: Roles of drosophila proto-oncogenes and growth factor homologs during development of the fly; Interaction of oncogenes with differentiation programs; Genetics of src: structure and functional organization of a protein tyrosine kinase; Structures and activities of activated abl oncogenes; Eukaryotic RAS proteins and yeast proteins with which they interact. This book presents up-to-data review articles on oncogenes. The editor includes five contributions which critically evaluate recent research in the field.

  2. Oncogenic Ras influences the expression of multiple lncRNAs.

    PubMed

    Kotake, Yojiro; Naemura, Madoka; Kitagawa, Kyoko; Niida, Hiroyuki; Tsunoda, Toshiyuki; Shirasawa, Senji; Kitagawa, Masatoshi

    2016-08-01

    Recent ultrahigh-density tiling array and large-scale transcriptome analysis have revealed that large numbers of long non-coding RNAs (lncRNAs) are transcribed in mammals. Several lncRNAs have been implicated in transcriptional regulation, organization of nuclear structure, and post-transcriptional processing. However, the regulation of expression of lncRNAs is less well understood. Here, we show that the exogenous and endogenous expression of an oncogenic form of small GTPase Ras (called oncogenic Ras) decrease the expression of lncRNA ANRIL (antisense non-coding RNA in the INK4 locus), which is involved in the regulation of cellular senescence. We also show that forced expression of oncogenic Ras increases the expression of lncRNA PANDA (p21 associated ncRNA DNA damage activated), which is involved in the regulation of apoptosis. Microarray analysis demonstrated that expression of multiple lncRNAs fluctuated by forced expression of oncogenic Ras. These findings indicate that oncogenic Ras regulates the expression of a large number of lncRNAs including functional lncRNAs, such as ANRIL and PANDA. PMID:25501747

  3. HER-2/neu oncogene expression and proliferation in breast cancers.

    PubMed Central

    Bacus, S. S.; Ruby, S. G.; Weinberg, D. S.; Chin, D.; Ortiz, R.; Bacus, J. W.

    1990-01-01

    Amplification of the HER-2/neu proto-oncogene in breast cancer has been reported to correlate with poor patient prognosis. The proliferation, or growth fraction, of cells has also been shown to be of prognostic importance in breast cancer. A study was conducted to evaluate the correlation between HER-2/neu gene expression and proliferation in breast cancer. Quantitative immunohistochemical methods for the detection of the HER-2/neu protein expression and for assessing the proliferation fraction on frozen sections of tumor cells were used. The detection of epidermal growth factor receptor (EGFR) along with quantitative DNA ploidy analysis, also was performed on the same breast cancers. The results indicated two subgroups of invasive ductal carcinoma; 1) HER-2/neu overexpressing cases that were negative for EGFR expression and had low proliferation fraction, and a tetraploid DNA pattern (22 cases), and 2) other combinations of HER-2/neu expression and EGFR expression, with a high proliferation fraction and an aneuploid DNA pattern (38 cases). Eight cases of carcinoma in situ were positive for HER-2/neu overexpression and negative for EGFR expression, and had a high proliferation fraction and a tetraploid DNA pattern. Twenty-six cases of low-grade carcinoma exhibited low proliferation and a diploid DNA pattern. Images Figure 1 Figure 2 PMID:1973597

  4. Stromal control of oncogenic traits expressed in response to the overexpression of GLI2, a pleiotropic oncogene.

    PubMed

    Snijders, A M; Huey, B; Connelly, S T; Roy, R; Jordan, R C K; Schmidt, B L; Albertson, D G

    2009-02-01

    Hedgehog signaling is often activated in tumors, yet it remains unclear how GLI2, a transcription factor activated by this pathway, acts as an oncogene. We show that GLI2 is a pleiotropic oncogene. The overexpression induces genomic instability and blocks differentiation, likely mediated in part by enhanced expression of the stem cell gene SOX2. GLI2 also induces transforming growth factor (TGF)B1-dependent transdifferentiation of foreskin and tongue, but not gingival fibroblasts into myofibroblasts, creating an environment permissive for invasion by keratinocytes, which are in various stages of differentiation having downregulated GLI2. Thus, upregulated GLI2 expression is sufficient to induce a number of the acquired characteristics of tumor cells; however, the stroma, in a tissue-specific manner, determines whether certain GLI2 oncogenic traits are expressed. PMID:19015636

  5. Loss of oncogenic ras expression does not correlate with loss of tumorigenicity in human cells.

    PubMed Central

    Plattner, R; Anderson, M J; Sato, K Y; Fasching, C L; Der, C J; Stanbridge, E J

    1996-01-01

    ras oncogenes are mutated in at variety of human tumors, which suggests that they play an important role in human carcinogenesis. To determine whether continued oncogenic ras expression is necessary to maintain the malignant phenotype, we studied the human fibrosarcoma cell line, HT1080, which contains one mutated and one wild-type N-ras allele. We isolated a variant of this cell line that no longer contained the mutated copy of the N-ras gene. Loss of mutant N-ras resulted in cells that displayed a less transformed phenotype characterized by a flat morphology, decreased growth rate, organized actin stress fibers, and loss of anchorage-independent growth. The transformed phenotype was restored following reintroduction of mutant N-ras. Although loss of the oncogenic N-ras drastically affected in vitro growth parameters, the variant remained tumorigenic in nude mice indicating that mutated N-ras expression is not necessary for maintenance of the tumorigenic phenotype. We confirmed this latter observation in colon carcinoma cell lines that have lost activated K-ras expression via targeted knockout of the mutant K-ras gene. Images Fig. 1 Fig. 2 Fig. 3 Fig. 5 PMID:8692875

  6. The proto-oncogene c-ets is preferentially expressed in lymphoid cells.

    PubMed Central

    Chen, J H

    1985-01-01

    The transforming sequences of the avian acute leukemia virus, E26, contain two distinct oncogenes, v-mybE and v-ets, fused together. By using a probe containing v-ets sequences, polyadenylated transcripts of the c-ets proto-oncogene were detected in avian tissues; they included a major 7.0-kilobase and a minor 2.0-kilobase species. These c-ets mRNAs were detected at high levels only in lymphoid organs and in avian T and B lymphoid cell lines. A similar pattern of c-ets transcription was observed in human hematopoietic cell lines, with transcripts detected in lymphoid B and T cells but not in erythroid or myeloid cells. The E26 oncogene was inserted into an inducible expression vector, and a 90-kilodalton protein (bp90) was produced in bacteria. Rabbit antisera raised to purified bp90 precipitated P135gag-mybE-ets, the v-mybE-ets polyprotein expressed in E26-transformed cells, and also reacted with p50v-mybA, the transforming protein of the avian myeloblastosis virus. Antiserum to bp90 was absorbed with a bacterially synthesized v-mybA protein to remove anti-myb activity. The absorbed anti-bp90 serum retained the ability to immunoprecipitate P135gag-mybE-ets from E26-transformed cells and specifically reacted with a 56-kilodalton polypeptide (p56) detected in chicken lymphoid organs and in T and B lymphocytes of both avian and human origin. The data suggest that p56 is a translational product of the c-ets proto-oncogene and imply that p56 may be involved in regulating the growth of lymphoid cells. Images PMID:3018492

  7. Protein kinase Cι expression and oncogenic signaling mechanisms in cancer.

    PubMed

    Murray, Nicole R; Kalari, Krishna R; Fields, Alan P

    2011-04-01

    Accumulating evidence demonstrates that PKCι is an oncogene and prognostic marker that is frequently targeted for genetic alteration in many major forms of human cancer. Functional data demonstrate that PKCι is required for the transformed phenotype of lung, pancreatic, ovarian, prostate, colon, and brain cancer cells. Future studies will be required to determine whether PKCι is also an oncogene in the many other cancer types that also overexpress PKCι. Studies of PKCι using genetically defined models of tumorigenesis have revealed a critical role for PKCι in multiple stages of tumorigenesis, including tumor initiation, progression, and metastasis. Recent studies in a genetic model of lung adenocarcinoma suggest a role for PKCι in transformation of lung cancer stem cells. These studies have important implications for the therapeutic use of aurothiomalate (ATM), a highly selective PKCι signaling inhibitor currently undergoing clinical evaluation. Significant progress has been made in determining the molecular mechanisms by which PKCι drives the transformed phenotype, particularly the central role played by the oncogenic PKCι-Par6 complex in transformed growth and invasion, and of several PKCι-dependent survival pathways in chemo-resistance. Future studies will be required to determine the composition and dynamics of the PKCι-Par6 complex, and the mechanisms by which oncogenic signaling through this complex is regulated. Likewise, a better understanding of the critical downstream effectors of PKCι in various human tumor types holds promise for identifying novel prognostic and surrogate markers of oncogenic PKCι activity that may be clinically useful in ongoing clinical trials of ATM. PMID:20945390

  8. Derepression of hTERT gene expression promotes escape from oncogene-induced cellular senescence

    PubMed Central

    Patel, Priyanka L.; Suram, Anitha; Mirani, Neena; Bischof, Oliver; Herbig, Utz

    2016-01-01

    Oncogene-induced senescence (OIS) is a critical tumor-suppressing mechanism that restrains cancer progression at premalignant stages, in part by causing telomere dysfunction. Currently it is unknown whether this proliferative arrest presents a stable and therefore irreversible barrier to cancer progression. Here we demonstrate that cells frequently escape OIS induced by oncogenic H-Ras and B-Raf, after a prolonged period in the senescence arrested state. Cells that had escaped senescence displayed high oncogene expression levels, retained functional DNA damage responses, and acquired chromatin changes that promoted c-Myc–dependent expression of the human telomerase reverse transcriptase gene (hTERT). Telomerase was able to resolve existing telomeric DNA damage response foci and suppressed formation of new ones that were generated as a consequence of DNA replication stress and oncogenic signals. Inhibition of MAP kinase signaling, suppressing c-Myc expression, or inhibiting telomerase activity, caused telomere dysfunction and proliferative defects in cells that had escaped senescence, whereas ectopic expression of hTERT facilitated OIS escape. In human early neoplastic skin and breast tissue, hTERT expression was detected in cells that displayed features of senescence, suggesting that reactivation of telomerase expression in senescent cells is an early event during cancer progression in humans. Together, our data demonstrate that cells arrested in OIS retain the potential to escape senescence by mechanisms that involve derepression of hTERT expression. PMID:27503890

  9. Derepression of hTERT gene expression promotes escape from oncogene-induced cellular senescence.

    PubMed

    Patel, Priyanka L; Suram, Anitha; Mirani, Neena; Bischof, Oliver; Herbig, Utz

    2016-08-23

    Oncogene-induced senescence (OIS) is a critical tumor-suppressing mechanism that restrains cancer progression at premalignant stages, in part by causing telomere dysfunction. Currently it is unknown whether this proliferative arrest presents a stable and therefore irreversible barrier to cancer progression. Here we demonstrate that cells frequently escape OIS induced by oncogenic H-Ras and B-Raf, after a prolonged period in the senescence arrested state. Cells that had escaped senescence displayed high oncogene expression levels, retained functional DNA damage responses, and acquired chromatin changes that promoted c-Myc-dependent expression of the human telomerase reverse transcriptase gene (hTERT). Telomerase was able to resolve existing telomeric DNA damage response foci and suppressed formation of new ones that were generated as a consequence of DNA replication stress and oncogenic signals. Inhibition of MAP kinase signaling, suppressing c-Myc expression, or inhibiting telomerase activity, caused telomere dysfunction and proliferative defects in cells that had escaped senescence, whereas ectopic expression of hTERT facilitated OIS escape. In human early neoplastic skin and breast tissue, hTERT expression was detected in cells that displayed features of senescence, suggesting that reactivation of telomerase expression in senescent cells is an early event during cancer progression in humans. Together, our data demonstrate that cells arrested in OIS retain the potential to escape senescence by mechanisms that involve derepression of hTERT expression. PMID:27503890

  10. Cancer induction by restriction of oncogene expression to the stem cell compartment

    PubMed Central

    Pérez-Caro, María; Cobaleda, César; González-Herrero, Inés; Vicente-Dueñas, Carolina; Bermejo-Rodríguez, Camino; Sánchez-Beato, Margarita; Orfao, Alberto; Pintado, Belén; Flores, Teresa; Sánchez-Martín, Manuel; Jiménez, Rafael; Piris, Miguel A; Sánchez-García, Isidro

    2009-01-01

    In human cancers, all cancerous cells carry the oncogenic genetic lesions. However, to elucidate whether cancer is a stem cell-driven tissue, we have developed a strategy to limit oncogene expression to the stem cell compartment in a transgenic mouse setting. Here, we focus on the effects of the BCR-ABLp210 oncogene, associated with chronic myeloid leukaemia (CML) in humans. We show that CML phenotype and biology can be established in mice by restricting BCR-ABLp210 expression to stem cell antigen 1 (Sca1)+ cells. The course of the disease in Sca1-BCR-ABLp210 mice was not modified on STI571 treatment. However, BCR-ABLp210-induced CML is reversible through the unique elimination of the cancer stem cells (CSCs). Overall, our data show that oncogene expression in Sca1+ cells is all that is required to fully reprogramme it, giving rise to a full-blown, oncogene-specified tumour with all its mature cellular diversity, and that elimination of the CSCs is enough to eradicate the whole tumour. PMID:19037256

  11. Somatic Copy Number Alterations at Oncogenic Loci Show Diverse Correlations with Gene Expression

    PubMed Central

    Roszik, Jason; Wu, Chang-Jiun; Siroy, Alan E.; Lazar, Alexander J.; Davies, Michael A; Woodman, Scott E; Kwong, Lawrence N

    2016-01-01

    Somatic copy number alterations (SCNAs) affecting oncogenic drivers have a firmly established role in promoting cancer. However, no agreed-upon standard exists for calling locus-specific amplifications and deletions in each patient sample. Here, we report the correlative analysis of copy number amplitude and length with gene expression across 6,109 samples from The Cancer Genome Atlas (TCGA) dataset across 16 cancer types. Using specificity, sensitivity, and precision-based scores, we assigned optimized amplitude and length cutoffs for nine recurrent SCNAs affecting known oncogenic drivers, using mRNA expression as a functional readout. These cutoffs captured the majority of SCNA-driven, highly-expression-altered samples. The majority of oncogenes required only amplitude cutoffs, as high amplitude samples were almost invariably focal; however, CDKN2A and PTEN uniquely required both amplitude and length cutoffs as primary predictors. For PTEN, these extended to downstream AKT activation. In contrast, SCNA genes located peri-telomerically or in fragile sites showed poor expression-copy number correlations. Overall, our analyses identify optimized amplitude and length cutoffs as efficient predictors of gene expression changes for specific oncogenic SCNAs, yet warn against one-size-fits-all interpretations across all loci. Our results have implications for cancer data analyses and the clinic, where copy number and mutation data are increasingly used to personalize cancer therapy. PMID:26787600

  12. Somatic Copy Number Alterations at Oncogenic Loci Show Diverse Correlations with Gene Expression

    NASA Astrophysics Data System (ADS)

    Roszik, Jason; Wu, Chang-Jiun; Siroy, Alan E.; Lazar, Alexander J.; Davies, Michael A.; Woodman, Scott E.; Kwong, Lawrence N.

    2016-01-01

    Somatic copy number alterations (SCNAs) affecting oncogenic drivers have a firmly established role in promoting cancer. However, no agreed-upon standard exists for calling locus-specific amplifications and deletions in each patient sample. Here, we report the correlative analysis of copy number amplitude and length with gene expression across 6,109 samples from The Cancer Genome Atlas (TCGA) dataset across 16 cancer types. Using specificity, sensitivity, and precision-based scores, we assigned optimized amplitude and length cutoffs for nine recurrent SCNAs affecting known oncogenic drivers, using mRNA expression as a functional readout. These cutoffs captured the majority of SCNA-driven, highly-expression-altered samples. The majority of oncogenes required only amplitude cutoffs, as high amplitude samples were almost invariably focal; however, CDKN2A and PTEN uniquely required both amplitude and length cutoffs as primary predictors. For PTEN, these extended to downstream AKT activation. In contrast, SCNA genes located peri-telomerically or in fragile sites showed poor expression-copy number correlations. Overall, our analyses identify optimized amplitude and length cutoffs as efficient predictors of gene expression changes for specific oncogenic SCNAs, yet warn against one-size-fits-all interpretations across all loci. Our results have implications for cancer data analyses and the clinic, where copy number and mutation data are increasingly used to personalize cancer therapy.

  13. The MYC 3' Wnt-Responsive Element Drives Oncogenic MYC Expression in Human Colorectal Cancer Cells.

    PubMed

    Rennoll, Sherri A; Eshelman, Melanie A; Raup-Konsavage, Wesley M; Kawasawa, Yuka Imamura; Yochum, Gregory S

    2016-01-01

    Mutations in components of the Wnt/β-catenin signaling pathway drive colorectal cancer (CRC) by deregulating expression of downstream target genes including the c-MYC proto-oncogene (MYC). The critical regulatory DNA enhancer elements that control oncogenic MYC expression in CRC have yet to be fully elucidated. In previous reports, we correlated T-cell factor (TCF) and β-catenin binding to the MYC 3' Wnt responsive DNA element (MYC 3' WRE) with MYC expression in HCT116 cells. Here we used CRISPR/Cas9 to determine whether this element is a critical driver of MYC. We isolated a clonal population of cells that contained a deletion of a single TCF binding element (TBE) within the MYC 3' WRE. This deletion reduced TCF/β-catenin binding to this regulatory element and decreased MYC expression. Using RNA-Seq analysis, we found altered expression of genes that regulate metabolic processes, many of which are known MYC target genes. We found that 3' WRE-Mut cells displayed a reduced proliferative capacity, diminished clonogenic growth, and a decreased potential to form tumors in vivo. These findings indicate that the MYC 3' WRE is a critical driver of oncogenic MYC expression and suggest that this element may serve as a therapeutic target for CRC. PMID:27223305

  14. Tissue-specific expression and developmental regulation of the human fgr proto-oncogene.

    PubMed Central

    Ley, T J; Connolly, N L; Katamine, S; Cheah, M S; Senior, R M; Robbins, K C

    1989-01-01

    In this study, we show that c-fgr proto-oncogene expression is limited to normal peripheral blood granulocytes, monocytes, and alveolar macrophages, all of which contain 50 to 100 copies of c-fgr mRNA per cell. The c-fgr RNA molecules in these cells consisted of partially spliced transcripts containing intron 7 and completely spliced molecules capable of encoding the predicted p55 c-fgr protein. The splicing of intron 7 appeared to occur after the splicing of most of the other introns; partially spliced molecules containing intron 7 did not appear to be transported into the cytoplasm. Very low levels of fgr transcripts were also present in U937 promonocytic cells and increased in abundance with 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced differentiation. The level of fgr transcripts began to increase 2 to 4 h after TPA addition, peaked at 8 h, and subsequently declined. Since we found that the half-life of fgr mRNA was longer than 8 h, these changes are best explained by transient transcriptional activation of fgr during TPA-induced differentiation, although nuclear runoff experiments were not sensitive enough to detect this event. Cycloheximide also caused accumulation of c-fgr transcripts in U937 cells; no superinduction was observed when TPA and cycloheximide were added at the same time. Induction by either agent was blocked with actinomycin D. These results demonstrate that the c-fgr gene is expressed in a tissue- and development-specific fashion and suggest that constitutive expression of c-fgr in U937 cells is regulated by a labile transcriptional repressor. Images PMID:2538725

  15. Proto-oncogene expression in porcine myocardium subjected to ischemia and reperfusion.

    PubMed

    Brand, T; Sharma, H S; Fleischmann, K E; Duncker, D J; McFalls, E O; Verdouw, P D; Schaper, W

    1992-12-01

    The molecular basis of myocardial adaptation to ischemia and reperfusion is poorly understood. It is thought that nuclear proto-oncogenes act as third messengers, converting cytoplasmic signal transduction into long-term changes of gene expression. We studied the expression of six nuclear proto-oncogenes (Egr-1, c-fos, fosB, c-jun, junB, and c-myc) in myocardium subjected to ischemia and reperfusion in anesthetized pigs. Stunning was achieved by two 10-minute left anterior descending coronary artery occlusions separated by 30 minutes of reperfusion. Hearts were excised after the first occlusion, after the first reperfusion, and at 30, 120, 150, and 210 minutes of reperfusion after the second occlusion. Total RNA was prepared from stunned as well as normally perfused myocardial tissue and subjected to Northern blotting. The response of the six nuclear proto-oncogenes varied.fosB gene expression was never detected. The c-myc gene was expressed, but its level was unchanged by ischemia. c-jun expression was slightly increased by ischemia (3.1 +/- 0.6-fold). The c-fos, Egr-1, and junB genes were highly induced, being fivefold to sevenfold higher in experimental than in control tissue. In three animals pretreated with the beta 1-antagonist metoprolol and then subjected to the above experimental protocol, the induction of proto-oncogenes was similar to that in nonblocked controls. Our results show that the myocardial adaptive response to ischemic stress includes the induction of at least four transcription factors that may be further operative in repair processes and angiogenesis. PMID:1385005

  16. The TPR-MET oncogenic rearrangement is present and expressed in human gastric carcinoma and precursor lesions.

    PubMed Central

    Soman, N R; Correa, P; Ruiz, B A; Wogan, G N

    1991-01-01

    The TPR-MET oncogenic rearrangement was originally observed in an in vitro transformed human osteosarcoma cell line. Recently, we detected the expression of this rearrangement at very low levels in several cell lines derived from human tumors of nonhematopoietic origin using a highly sensitive method based on polymerase chain reaction amplification of the transcript. We report here the results of analysis of TPR-MET expression in cell lines derived from human gastric tumors and 22 biopsy samples of human gastric mucosa showing cancer or precursor lesions. The rearranged RNA was expressed in all four cell lines as well as in biopsy samples from 12 of the 22 patients. Overexpression of TPR-MET RNA in superficial gastritis lesions with hyperplasia of glandular neck cells suggests the possible involvement of this oncogene at an early stage of gastric tumorigenesis. Analysis of gastric biopsy samples for RAS gene mutations showed base substitutions occurring in the codon 12 region of Ki- and Ha-RAS genes in four cases, including two precursor lesions. Images PMID:2052572

  17. Normal Expression of a Rearranged and Mutated c-myc Oncogene after Transfection into Fibroblasts

    NASA Astrophysics Data System (ADS)

    Richman, Adam; Hayday, Adrian

    1989-10-01

    Expression of the c-myc oncogene is deregulated in a variety of malignancies. Rearrangement and mutation of the c-myc locus is a characteristic feature of human Burkitt's lymphoma. Whether deregulation is solely a result of mutation of c-myc or whether it is influenced by the transformed B cell context has not been determined. A translocated and mutated allele of c-myc was stably transfected into fibroblasts. The rearranged allele was expressed indistinguishably from a normal c-myc gene: it had serum-regulated expression, was transcribed with normal promoter preference, and was strongly attenuated. Thus mutations by themselves are insufficient to deregulate c-myc transcription.

  18. Oncogene mRNA Imaging with Radionuclide-PNA-Peptides

    SciTech Connect

    Wickstrom, Eric

    2008-03-19

    New cancer gene hybridization probes to carry radionuclides were made. Noninvasive technetium-99m gamma imaging of CCND1 cancer gene activity in human breast cancer tumors in mice was demonstrated, followed by noninvasive technetium-99m imaging of MYC cancer gene activity. Noninvasive imaging of CCND1 cancer gene activity in human breast cancer tumors in mice was demonstrated with a positron-emitting copper-64 probe, followed by noninvasive positron imaging of IRS1 cancer gene activity.

  19. Cellular oncogene expression following exposure of mice to {gamma}-rays

    SciTech Connect

    Anderson, A.; Woloschak, G.E.

    1991-06-12

    We examined the effects of total body exposure of BCF1 mice to {gamma}-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 {gamma}-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.

  20. Differential effects on ARF stability by normal vs. oncogenic levels of c-Myc expression

    PubMed Central

    Chen, Delin; Kon, Ning; Zhong, Jiayun; Zhang, Pingzhao; Yu, Long; Gu, Wei

    2013-01-01

    SUMMARY ARF suppresses aberrant cell growth upon c-Myc overexpression through activating p53 responses. Nevertheless, the precise mechanism by which ARF specifically, restrains the oncogenic potential of c-Myc without affecting its normal physiological function is not well understood. Here, we show that low levels of c-Myc expression stimulate cell proliferation whereas high levels inhibit through activating the ARF-p53 response. Although the mRNA levels of ARF are induced under both scenarios, the accumulation of ARF protein occurs only when ULF-mediated degradation of ARF is inhibited by c-Myc overexpression. Moreover, the levels of ARF are reduced through ULF-mediated ubiquitination upon DNA damage. Blocking ARF degradation by c-Myc overexpression dramatically stimulates the apoptotic responses. Our study reveals that ARF stability control is crucial for differentiating normal (low) vs. oncogenic (high) levels of c-Myc expression and suggests that differential effects on ULF- mediated ARF ubiquitination by c-Myc levels act as a barrier in oncogene-induced stress responses. PMID:23747016

  1. Increased H+ efflux is sufficient to induce dysplasia and necessary for viability with oncogene expression

    PubMed Central

    Grillo-Hill, Bree K; Choi, Changhoon; Jimenez-Vidal, Maite; Barber, Diane L

    2015-01-01

    Intracellular pH (pHi) dynamics is increasingly recognized as an important regulator of a range of normal and pathological cell behaviors. Notably, increased pHi is now acknowledged as a conserved characteristic of cancers and in cell models is confirmed to increase proliferation and migration as well as limit apoptosis. However, the significance of increased pHi for cancer in vivo remains unresolved. Using Drosophila melanogaster, we show that increased pHi is sufficient to induce dysplasia in the absence of other transforming cues and potentiates growth and invasion with oncogenic Ras. Using a genetically encoded biosensor we also confirm increased pHi in situ. Moreover, in Drosophila models and clonal human mammary cells we show that limiting H+ efflux with oncogenic Raf or Ras induces acidosis and synthetic lethality. Further, we show lethality in invasive primary tumor cell lines with inhibiting H+ efflux. Synthetic lethality with reduced H+ efflux and activated oncogene expression could be exploited therapeutically to restrain cancer progression while limiting off-target effects. DOI: http://dx.doi.org/10.7554/eLife.03270.001 PMID:25793441

  2. Expression of BCR-ABL1 oncogene relative to ABL1 gene changes overtime in chronic myeloid leukemia

    SciTech Connect

    Gupta, Manu; Milani, Lili; Hermansson, Monica; Simonsson, Bengt; Markevaern, Berit; Syvaenen, Ann Christine; Barbany, Gisela

    2008-02-15

    Using a quantitative single nucleotide polymorphism (SNP) assay we have investigated the changes in the expression of the BCR-ABL1 oncogene relative to the wild-type ABL1 and BCR alleles in cells from chronic myeloid leukemia (CML) patients not responding to therapy. The results show a progressive increase in the BCR-ABL1 oncogene expression at the expense of decreased expression of the ABL1 allele, not involved in the fusion. No relative changes in the expression of the two BCR alleles were found. These results demonstrate that allele-specific changes in gene expression, with selective, progressive silencing of the wild-type ABL1 allele in favor of the oncogenic BCR-ABL1 allele occur in CML patients with therapy-resistant disease.

  3. STAT5 Outcompetes STAT3 To Regulate the Expression of the Oncogenic Transcriptional Modulator BCL6

    PubMed Central

    Walker, Sarah R.; Nelson, Erik A.; Yeh, Jennifer E.; Pinello, Luca; Yuan, Guo-Cheng

    2013-01-01

    Inappropriate activation of the transcription factors STAT3 and STAT5 has been shown to drive cancer pathogenesis through dysregulation of genes involved in cell survival, growth, and differentiation. Although STAT3 and STAT5 are structurally related, they can have opposite effects on key genes, including BCL6. BCL6, a transcriptional repressor, has been shown to be oncogenic in diffuse large B cell lymphoma. BCL6 also plays an important role in breast cancer pathogenesis, a disease in which STAT3 and STAT5 can be activated individually or concomitantly. To determine the mechanism by which these oncogenic transcription factors regulate BCL6 transcription, we analyzed their effects at the levels of chromatin and gene expression. We found that STAT3 increases expression of BCL6 and enhances recruitment of RNA polymerase II phosphorylated at a site associated with transcriptional initiation. STAT5, in contrast, represses BCL6 expression below basal levels and decreases the association of RNA polymerase II at the gene. Furthermore, the repression mediated by STAT5 is dominant over STAT3-mediated induction. STAT5 exerts this effect by displacing STAT3 from one of the two regulatory regions to which it binds. These findings may underlie the divergent biology of breast cancers containing activated STAT3 alone or in conjunction with activated STAT5. PMID:23716595

  4. Tocopherol Succinate: Modulation of Antioxidant Enzymes and Oncogene Expression, and Hematopoietic Recovery

    SciTech Connect

    Singh, Vijay K.; Parekh, Vaishali I.; Brown, Darren S.; Kao, Tzu-Cheg; Mog, Steven R.

    2011-02-01

    Purpose: A class of naturally occurring isoforms of tocopherol (tocols) was shown to have varying degrees of protection when administered before radiation exposure. We recently demonstrated that {alpha}-tocopherol succinate (TS) is a potential radiation prophylactic agent. Our objective in this study was to further investigate the mechanism of action of TS in mice exposed to {sup 60}Co {gamma}-radiation. Methods and Materials: We evaluated the effects of TS on expression of antioxidant enzymes and oncogenes by quantitative RT-PCR in bone marrow cells of {sup 60}Co {gamma}-irradiated mice. Further, we tested the ability of TS to rescue and repopulate hematopoietic stem cells by analyzing bone marrow cellularity and spleen colony forming unit in spleen of TS-injected and irradiated mice. Results: Our results demonstrate that TS modulated the expression of antioxidant enzymes and inhibited expression of oncogenes in irradiated mice at different time points. TS also increased colony forming unit-spleen numbers and bone marrow cellularity in irradiated mice. Conclusions: Results provide additional support for the observed radioprotective efficacy of TS and insight into mechanisms.

  5. AID-expressing epithelium is protected from oncogenic transformation by an NKG2D surveillance pathway.

    PubMed

    Pérez-García, Arantxa; Pérez-Durán, Pablo; Wossning, Thomas; Sernandez, Isora V; Mur, Sonia M; Cañamero, Marta; Real, Francisco X; Ramiro, Almudena R

    2015-10-01

    Activation-induced deaminase (AID) initiates secondary antibody diversification in germinal center B cells, giving rise to higher affinity antibodies through somatic hypermutation (SHM) or to isotype-switched antibodies through class switch recombination (CSR). SHM and CSR are triggered by AID-mediated deamination of cytosines in immunoglobulin genes. Importantly, AID activity in B cells is not restricted to Ig loci and can promote mutations and pro-lymphomagenic translocations, establishing a direct oncogenic mechanism for germinal center-derived neoplasias. AID is also expressed in response to inflammatory cues in epithelial cells, raising the possibility that AID mutagenic activity might drive carcinoma development. We directly tested this hypothesis by generating conditional knock-in mouse models for AID overexpression in colon and pancreas epithelium. AID overexpression alone was not sufficient to promote epithelial cell neoplasia in these tissues, in spite of displaying mutagenic and genotoxic activity. Instead, we found that heterologous AID expression in pancreas promotes the expression of NKG2D ligands, the recruitment of CD8(+) T cells, and the induction of epithelial cell death. Our results indicate that AID oncogenic potential in epithelial cells can be neutralized by immunosurveillance protective mechanisms. PMID:26282919

  6. Pervasive transcription read-through promotes aberrant expression of oncogenes and RNA chimeras in renal carcinoma

    PubMed Central

    Grosso, Ana R; Leite, Ana P; Carvalho, Sílvia; Matos, Mafalda R; Martins, Filipa B; Vítor, Alexandra C; Desterro, Joana MP; Carmo-Fonseca, Maria; de Almeida, Sérgio F

    2015-01-01

    Aberrant expression of cancer genes and non-canonical RNA species is a hallmark of cancer. However, the mechanisms driving such atypical gene expression programs are incompletely understood. Here, our transcriptional profiling of a cohort of 50 primary clear cell renal cell carcinoma (ccRCC) samples from The Cancer Genome Atlas (TCGA) reveals that transcription read-through beyond the termination site is a source of transcriptome diversity in cancer cells. Amongst the genes most frequently mutated in ccRCC, we identified SETD2 inactivation as a potent enhancer of transcription read-through. We further show that invasion of neighbouring genes and generation of RNA chimeras are functional outcomes of transcription read-through. We identified the BCL2 oncogene as one of such invaded genes and detected a novel chimera, the CTSC-RAB38, in 20% of ccRCC samples. Collectively, our data highlight a novel link between transcription read-through and aberrant expression of oncogenes and chimeric transcripts that is prevalent in cancer. DOI: http://dx.doi.org/10.7554/eLife.09214.001 PMID:26575290

  7. AID-expressing epithelium is protected from oncogenic transformation by an NKG2D surveillance pathway

    PubMed Central

    Pérez-García, Arantxa; Pérez-Durán, Pablo; Wossning, Thomas; Sernandez, Isora V; Mur, Sonia M; Cañamero, Marta; Real, Francisco X; Ramiro, Almudena R

    2015-01-01

    Activation-induced deaminase (AID) initiates secondary antibody diversification in germinal center B cells, giving rise to higher affinity antibodies through somatic hypermutation (SHM) or to isotype-switched antibodies through class switch recombination (CSR). SHM and CSR are triggered by AID-mediated deamination of cytosines in immunoglobulin genes. Importantly, AID activity in B cells is not restricted to Ig loci and can promote mutations and pro-lymphomagenic translocations, establishing a direct oncogenic mechanism for germinal center-derived neoplasias. AID is also expressed in response to inflammatory cues in epithelial cells, raising the possibility that AID mutagenic activity might drive carcinoma development. We directly tested this hypothesis by generating conditional knock-in mouse models for AID overexpression in colon and pancreas epithelium. AID overexpression alone was not sufficient to promote epithelial cell neoplasia in these tissues, in spite of displaying mutagenic and genotoxic activity. Instead, we found that heterologous AID expression in pancreas promotes the expression of NKG2D ligands, the recruitment of CD8+ T cells, and the induction of epithelial cell death. Our results indicate that AID oncogenic potential in epithelial cells can be neutralized by immunosurveillance protective mechanisms. PMID:26282919

  8. Kita Driven Expression of Oncogenic HRAS Leads to Early Onset and Highly Penetrant Melanoma in Zebrafish

    PubMed Central

    Santoriello, Cristina; Gennaro, Elisa; Anelli, Viviana; Distel, Martin; Kelly, Amanda; Köster, Reinhard W.; Hurlstone, Adam; Mione, Marina

    2010-01-01

    Background 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. Methodology and Principal Findings 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. Conclusions and Significance 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. PMID:21170325

  9. Oncogenic BRAFV600E inhibits BIM expression to promote melanoma cell survival

    PubMed Central

    Cartlidge, Robert A.; Thomas, G. R.; Cagnol, Sebastien; Jong, Kimberly A.; Molton, Sarah A.; Finch, Andrew J.; McMahon, Martin

    2016-01-01

    Summary Somatic activating mutations of BRAF are the earliest and most common genetic abnormality detected in the genesis of human melanoma. However, the mechanism(s) by which activated BRAF promotes melanoma cell cycle progression and/or survival remain unclear. Here we demonstrate that expression of BIM, a pro-apoptotic member of the BCL-2 family, is inhibited by BRAF → MEK → ERK signaling in mouse and human melanocytes and in human melanoma cells. Trophic factor deprivation of melanocytes leads to elevated BIM expression. However, re-addition of trophic factors or activation of a conditional form of BRAFV600E leads to rapid inhibition of BIM expression. In both cases, inhibition of BIM expression was dependent on the activity of MEK1/2 and the proteasome. Consistent with these observations, pharmacological inhibition of BRAFV600E or MEK1/2 in human melanoma cells (using PLX4720 and CI-1040 respectively) led to a striking elevation of BIM expression. Re-activation of BRAF → MEK → ERK signaling led to phosphorylation of BIM-EL on serine 69 and its subsequent degradation. Interestingly, endogenous expression of BIM in melanoma cells was insufficient to induce apoptosis unless combined with serum deprivation. Under these circumstances, inhibition of BIM expression by RNA interference provided partial protection from apoptosis. These data suggest that regulation of BIM expression by BRAF → MEK → ERK signaling is one mechanism by which oncogenic BRAFV600E can influence the aberrant physiology of melanoma cells. PMID:18715233

  10. SIRT6 promotes COX-2 expression and acts as an oncogene in skin cancer

    PubMed Central

    Ming, Mei; Han, Weinong; Zhao, Baozhong; Sundaresan, Nagalingam R.; Deng, Chu-Xia; Gupta, Mahesh; He, Yu-Ying

    2014-01-01

    SIRT6 is a SIR2 family member that regulates multiple molecular pathways involved in metabolism, genomic stability and aging. It has been proposed previously that SIRT6 is a tumor suppressor in cancer. Here we challenge this concept by presenting evidence that skin-specific deletion of SIRT6 in the mouse inhibits skin tumorigenesis. SIRT6 promoted expression of COX-2 by repressing AMPK signaling, thereby increasing cell proliferation and survival and in the skin epidermis. SIRT6 expression in skin keratinocytes was increased by exposure to UVB light through activation of the AKT pathway. Clinically, we found that SIRT6 was upregulated in human skin squamous cell carcinoma. Taken together, our results provide evidence that SIRT6 functions an oncogene in the epidermis and suggest greater complexity to its role in epithelial carcinogenesis. PMID:25320180

  11. Oncogenic relevant defensins: expression pattern and proliferation characteristics of human tumor cell lines.

    PubMed

    Winter, Jochen; Kraus, Dominik; Reckenbeil, Jan; Probstmeier, Rainer

    2016-06-01

    The objective of this study was to investigate gene expression levels of oncogenic relevant human defensins and their impact on proliferation rates of 29 cell lines derived from main types of different tumor origins. Differential gene expression analysis of human defensins was performed by real-time PCR experiments. The proliferation rate of tumor cells that had been cultivated in the absence or presence of biologically active peptides was analyzed with a lactate dehydrogenase assay kit. At least one member of the defensin family was expressed in each tumor cell line, whereby α-defensin (DEFA1), DEFA2, or DEFA3 transcripts could be ubiquitously detected. Cell lines of neural origin (glioma, neuroblastoma, and small-cell lung carcinoma) expressed far less human β-defensins (hBDs) in comparison to other tumor types. The expression level of a specific defensin in various cell lines could vary by more than five orders of magnitude. Compensatory mechanisms on the expression levels of the different defensins could not be strictly observed. Only in 3 out of 29 tumor cell lines the proliferation rate was affected after defensin stimulation. The variable appearance of defensins, as well as the cell line-restricted functional activity, argues for the integration of defensins in complex cellular and molecular networks that tolerate rather flexible expression patterns. PMID:26711780

  12. Amplification and expression of the c-myc oncogene in human lung cancer cell lines.

    PubMed

    Little, C D; Nau, M M; Carney, D N; Gazdar, A F; Minna, J D

    Genetic changes involving the c-myc oncogene have been observed in human tumours. In particular, the c-myc gene is translocated in Burkitt's lymphoma and is amplified in the human promyelocytic leukaemia cell line, HL-60, which contains double minute chromosomes (DMs). More recently, an amplified c-myc gene has been positioned on a chromosomal homogeneous staining region (HSR) in a human colon cancer cell line, COLO 320, with neuroendocrine properties. Furthermore, c-myc is expressed in increased amounts in some human tumour lines, and in some cases, human small cell lung cancers (SCLC) contain DMs and HSRs. These findings prompted us to study the c-myc gene and its RNA expression in a series of human lung cancer cell lines. We now report amplification and expression of the c-myc oncogene in a system other than B-cell lymphomas, namely human lung cancer. Of 18 human lung cancer cell lines tested, 8 showed an amplified 12.5-kilobase (kb) EcoRI c-myc DNA band. Of particular interest are five SCLC lines with a high degree of c-myc DNA amplification (20-76-fold) and greatly increased levels of c-myc RNA. All five lines reside in the variant class of SCLC (SCLC-V) characterized by altered morphology, lack of expression of some SCLC-differentiated functions and more malignant behaviour than pure SCLC. Three of the five lines which have been karyotyped also contain DMs or HSRs. The finding of a greatly amplified c-myc gene in all cell lines of the SCLC-V class examined strongly suggests a role for the c-myc gene in the phenotypic conversion and malignant behaviour of human lung cancer. PMID:6646201

  13. Enhanced human papillomavirus type 8 oncogene expression levels are crucial for skin tumorigenesis in transgenic mice

    SciTech Connect

    Hufbauer, M.; Lazic, D.; Akguel, B.; Brandsma, J.L.; Pfister, H.; Weissenborn, S.J.

    2010-08-01

    Human papillomavirus 8 (HPV8) is involved in skin cancer development in epidermodysplasia verruciformis patients. Transgenic mice expressing HPV8 early genes (HPV8-CER) developed papillomas, dysplasias and squamous cell carcinomas. UVA/B-irradiation and mechanical wounding of HPV8-CER mouse skin led to prompt papilloma induction in about 3 weeks. The aim of this study was to analyze the kinetics and level of transgene expression in response to skin irritations. Transgene expression was already enhanced 1 to 2 days after UVA/B-irradiation or tape-stripping and maintained during papilloma development. The enhanced transgene expression could be assigned to UVB and not to UVA. Papilloma development was thus always paralleled by an increased transgene expression irrespective of the type of skin irritation. A knock-down of E6 mRNA by tattooing HPV8-E6-specific siRNA led to a delay and a lower incidence of papilloma development. This indicates that the early increase of viral oncogene expression is crucial for induction of papillomatosis.

  14. Expression of the Pokemon proto-oncogene in nasopharyngeal carcinoma cell lines and tissues.

    PubMed

    Jiao, Wei; Liu, Fei; Tang, Feng-Zhu; Lan, Jiao; Xiao, Rui-Ping; Chen, Xing-Zhou; Ye, Hui-Lan; Cai, Yong-Lin

    2013-01-01

    To study the differentiated expression of the proto-oncogene Pokemon in nasopharyngeal carcinoma (NPC) cell lines and tissues, mRNA and protein expression levels of CNE1, CNE2, CNE3 and C666-1 were detected separately by reverse transcription polymerase chain reaction (RT-PCR), real-time PCR and Western-blotting. The immortalized nasopharyngeal epithelial cell line NP69 was used as a control. The Pokemon protein expression level in biopsy specimens from chronic rhinitis patients and undifferentiated non keratinizing NPC patients was determined by Western-blotting and arranged from high to low: C666-1>CNE1>CNE2> CNE3>NP69. The Pokemon mRNA expression level was also arranged from high to low: CNE1>CNE2>NP69>C666-1>CNE3. Pokemon expression of NP69 and C666-1 obviously varied from mRNA to protein. The Pokemon protein level of NPC biopsy specimens was obviously higher than in chronic rhinitis. The data suggest that high Pokemon protein expression is closely associated with undifferentiated non-keratinizing NPC and may provide useful information for NPC molecular target therapy. PMID:24377524

  15. Arsenic trioxide inhibits cell proliferation and human papillomavirus oncogene expression in cervical cancer cells

    SciTech Connect

    Wang, Hongtao; Gao, Peng; Zheng, Jie

    2014-09-05

    Highlights: • As{sub 2}O{sub 3} inhibits growth of cervical cancer cells and expression of HPV oncogenes in these cells. • HPV-negative cervical cancer cells are more sensitive to As{sub 2}O{sub 3} than HPV-positive cervical cancer cells. • HPV-18 positive cervical cancer cells are more sensitive to As{sub 2}O{sub 3} than HPV-16 positive cancer cells. • Down-regulation of HPV oncogenes by As{sub 2}O{sub 3} is partially due to the diminished AP-1 binding. - Abstract: Arsenic trioxide (As{sub 2}O{sub 3}) has shown therapeutic effects in some leukemias and solid cancers. However, the molecular mechanisms of its anticancer efficacy have not been clearly elucidated, particularly in solid cancers. Our previous data showed that As{sub 2}O{sub 3} induced apoptosis of human papillomavirus (HPV) 16 DNA-immortalized human cervical epithelial cells and cervical cancer cells and inhibited the expression of HPV oncogenes in these cells. In the present study, we systemically examined the effects of As{sub 2}O{sub 3} on five human cervical cancer cell lines and explored the possible molecular mechanisms. MTT assay showed that HPV-negative C33A cells were more sensitive to growth inhibition induced by As{sub 2}O{sub 3} than HPV-positive cervical cancer cells, and HPV 18-positive HeLa and C4-I cells were more sensitive to As{sub 2}O{sub 3} than HPV 16-positive CaSki and SiHa cells. After As{sub 2}O{sub 3} treatment, both mRNA and protein levels of HPV E6 and E7 obviously decreased in all HPV positive cell lines. In contrast, p53 and Rb protein levels increased in all tested cell lines. Transcription factor AP-1 protein expression decreased significantly in HeLa, CaSki and C33A cells with ELISA method. These results suggest that As{sub 2}O{sub 3} is a potential anticancer drug for cervical cancer.

  16. The oncogenic transcription factor c-Jun regulates glutaminase expression and sensitizes cells to glutaminase-targeted therapy

    PubMed Central

    Lukey, Michael J.; Greene, Kai Su; Erickson, Jon W.; Wilson, Kristin F.; Cerione, Richard A.

    2016-01-01

    Many transformed cells exhibit altered glucose metabolism and increased utilization of glutamine for anabolic and bioenergetic processes. These metabolic adaptations, which accompany tumorigenesis, are driven by oncogenic signals. Here we report that the transcription factor c-Jun, product of the proto-oncogene JUN, is a key regulator of mitochondrial glutaminase (GLS) levels. Activation of c-Jun downstream of oncogenic Rho GTPase signalling leads to elevated GLS gene expression and glutaminase activity. In human breast cancer cells, GLS protein levels and sensitivity to GLS inhibition correlate strongly with c-Jun levels. We show that c-Jun directly binds to the GLS promoter region, and is sufficient to increase gene expression. Furthermore, ectopic overexpression of c-Jun renders breast cancer cells dependent on GLS activity. These findings reveal a role for c-Jun as a driver of cancer cell metabolic reprogramming, and suggest that cancers overexpressing JUN may be especially sensitive to GLS-targeted therapies. PMID:27089238

  17. H-ras oncogene expression and angiogenesis in experimental liver cirrhosis.

    PubMed

    Elpek, Gülsüm Özlem; Unal, Betül; Bozova, Sevgi

    2013-01-01

    Background. Proto-oncogenes, particularly ras, may not only affect cell proliferation but also contribute to angiogenesis by influencing both proangiogenic and antiangiogenic mediators. The aim of this study was to investigate whether any relationship exists between ras expression and angiogenesis during diethylnitrosamine- (DEN-) induced experimental liver fibrosis. Materials and Methods. Liver cirrhosis was induced in rats by intraperitoneal injections of DEN. The animals were sacrificed 2 weeks after the last administrations and a hepatectomy was performed. Masson's trichrome staining was used in the evaluation of the extent of liver fibrosis. The vascular density in portal and periportal areas was assessed by determining the count of CD34 labeled vessel sections. For quantitative evaluation of H-ras expression, in each section positive and negative cells were counted. Results. In fibrotic group H-ras expression was higher than that in nonfibrotic group and was more widespread in cirrhotic livers. Friedman's test showed that there was a significant correlation between H-ras expression and VD (P < 0.01). Conclusion. The results of this descriptive study reveal that H-ras expression gradually increases according to the severity of fibrosis and strongly correlates with angiogenesis. PMID:24235967

  18. Role of Cdc6 in re-replication in cells expressing human papillomavirus E7 oncogene.

    PubMed

    Fan, Xueli; Zhou, Yunying; Chen, Jason J

    2016-08-01

    The E7 oncoprotein of high-risk human papillomavirus (HPV) types induces DNA re-replication that contributes to carcinogenesis; however, the mechanism is not fully understood. To better understand the mechanism by which E7 induces re-replication, we investigated the expression and function of cell division cycle 6 (Cdc6) in E7-expressing cells. Cdc6 is a DNA replication initiation factor and exhibits oncogenic activities when overexpressed. We found that in E7-expressing cells, the steady-state level of Cdc6 protein was upregulated and its half-life was increased. Cdc6 was localized to the nucleus and associated with chromatin, especially upon DNA damage. Importantly, downregulation of Cdc6 reduced E7-induced re-replication. Interestingly, the level of Cdc6 phosphorylation at serine 54 (S54P) was increased in E7-expressing cells. S54P was associated with an increase in the total amount of Cdc6 and chromatin-bound Cdc6. DNA damage-enhanced upregulation and chromatin binding of Cdc6 appeared to be due to downregulation of cyclin-dependent kinase 1 (Cdk1) as Cdk1 knockdown increased Cdc6 levels. Furthermore, Cdk1 knockdown or inhibition led to re-replication. These findings shed light on the mechanism by which HPV induces genomic instability and may help identify potential targets for drug development. PMID:27207654

  19. The prognostic potential and oncogenic effects of PRR11 expression in hilar cholangiocarcinoma

    PubMed Central

    Qian, Baohua; Yu, Wenlong; Li, Wenfeng; Yu, Guanzhen; Gao, Yong

    2015-01-01

    PRR11 is a newly identified oncogene in lung cancer, yet its role in others tumors remains unclear. Gastrointestinal tissue microarrays were used to evaluate PRR11 expression and its association with clinical outcome was analyzed in patients with hilar cholangiocarcinoma. Overexpression of PRR11 was observed in esophageal, gastric, pancreatic, colorectal, and hilar cholangiocarcinoma. Expression of PRR11 correlated with lymph node metastasis and CA199 level in two HC patient cohorts. After an R0 resection, a high level of PRR11 expression was found to be an independent indicator of recurrence (P = 0.001). In cell culture, PRR11 silencing resulted in decreased cellular proliferation, cell migration, tumor growth of QBC939 cells. Microarray analysis revealed that several genes involved in cell proliferation, cell adhesion, and cell migration were altered in PRR11-knockout cells, including: vimentin (VIM), Ubiquitin carboxyl-terminal hydrolase 1 (UCHL1), early growth response protein (EGR1), and System A amino acid transporter1 (SNAT1). Silencing PRR11 inhibited the expression of UCHL1, EGR1, and SNAT1 proteins, with immunoassays revealing a significant correlation among the levels of these four proteins. These results indicate that PRR11 is an independent prognostic indicator for patients with HC. PMID:25971332

  20. Activation and repression by oncogenic MYC shape tumour-specific gene expression profiles.

    PubMed

    Walz, Susanne; Lorenzin, Francesca; Morton, Jennifer; Wiese, Katrin E; von Eyss, Björn; Herold, Steffi; Rycak, Lukas; Dumay-Odelot, Hélène; Karim, Saadia; Bartkuhn, Marek; Roels, Frederik; Wüstefeld, Torsten; Fischer, Matthias; Teichmann, Martin; Zender, Lars; Wei, Chia-Lin; Sansom, Owen; Wolf, Elmar; Eilers, Martin

    2014-07-24

    In mammalian cells, the MYC oncoprotein binds to thousands of promoters. During mitogenic stimulation of primary lymphocytes, MYC promotes an increase in the expression of virtually all genes. In contrast, MYC-driven tumour cells differ from normal cells in the expression of specific sets of up- and downregulated genes that have considerable prognostic value. To understand this discrepancy, we studied the consequences of inducible expression and depletion of MYC in human cells and murine tumour models. Changes in MYC levels activate and repress specific sets of direct target genes that are characteristic of MYC-transformed tumour cells. Three factors account for this specificity. First, the magnitude of response parallels the change in occupancy by MYC at each promoter. Functionally distinct classes of target genes differ in the E-box sequence bound by MYC, suggesting that different cellular responses to physiological and oncogenic MYC levels are controlled by promoter affinity. Second, MYC both positively and negatively affects transcription initiation independent of its effect on transcriptional elongation. Third, complex formation with MIZ1 (also known as ZBTB17) mediates repression of multiple target genes by MYC and the ratio of MYC and MIZ1 bound to each promoter correlates with the direction of response. PMID:25043018

  1. Posttranscriptional regulation of cellular gene expression by the c-myc oncogene

    SciTech Connect

    Prendergast, G.C.; Cole, M.D. . Dept. of Biology)

    1989-01-01

    The c-myc oncogene has been implicated in the development of many different cancers, yet the mechanism by which the c-myc protein alters cellular growth control has proven elusive. The authors used a cDNA hybridization difference assay to isolate two genes, mr1 and mr2, that were constitutively expressed (i.e., deregulated) in rodent fibroblast cell lines immortalized by transfection of a viral promoter-linked c-myc gene. Both cDNAs were serum inducible in quiescent G/sub o/ fibroblasts, suggesting that they are functionally related to cellular proliferative processes. Although there were significant differences in cytoplasmic mRNA levels between myc-immortalized and control cells, the rates of transcription and mRNA turnover of both genes were similar, suggesting that c-myc regulates mr1 and mr2 expression by some nuclear posttranscriptional mechanism. Their results provide evidence that c-myc can rapidly modulate cellular gene expression and suggest that c-myc may function in gene regulation at the level of RNA export, splicing, or nuclear RNA turnover.

  2. Effects of telomerase and viral oncogene expression on the in vitro growth of human chondrocytes.

    PubMed

    Martin, James A; Mitchell, Calista J; Klingelhutz, Aloysius J; Buckwalter, Joseph A

    2002-02-01

    Senescent chondrocytes accumulate with aging in articular cartilage, a process that interferes with cartilage homeostasis and increases the risk of cartilage degeneration. We showed previously that chondrocyte telomere length declines with donor age, which suggests that the aging process is telomere dependent. From these results we hypothesized that telomerase should delay the onset of senescence in cultured chondrocytes. Population doubling limits (PDL) were determined for chondrocytes expressing telomerase. We found that telomerase alone did not extend PDL beyond controls that senesced after 25 population doublings. The human papillomavirus 16 oncogenes E6 and E7 were transduced into the same cell population to investigate this telomere-independent form of senescence further. Chondrocytes expressing E6 and E7 grew longer than the telomerase cDNA (hTERT) cells but still senesced at 55 population doublings. In contrast, chondrocytes expressing telomerase with E6 and E7 grew vigorously past 100 population doublings. We conclude that although telomerase is necessary for the indefinite extension of chondrocyte life span, telomere-independent senescence limits PDL in vitro and may play a role in the age-related accumulation of senescent chondrocytes in vivo. PMID:11818423

  3. Tissue-specific expression and developmental regulation of the human fgr proto-oncogene

    SciTech Connect

    Levy, T.J. . Dept. of Medicine); Connolly, N.L.; Senior, R.M. ); Katamine, S.; Cheah, M.S.C.; Robbins

    1989-01-01

    In this study, the authors show that c-fgr proto-oncogene expression is limited to normal peripheral blood granulocytes, monocytes, and alveolar macrophages, all of which contain 50 to 100 copies of c-fgr mRNA per cell. The c-fgr RNA molecules in these cells consisted of partially spliced transcripts containing intron 7 and completely spliced molecules capable of encoding the predicted p55 c-fgr protein. The splicing of intron 7 appeared to occur after the splicing of most of the other introns; partially spliced molecules containing intron 7 did not appear to be transported into the cytoplasm. Very low levels of fgr transcripts were also present in U937 promonocytic cells and increased in abundance with 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced differentiation. The level of fgr transcripts began to increase 2 to 4 after TPA addition peaked at 8 h, and subsequently declined. Since the authors found that the half-life of fgr mRNA was longer than 8 h, these changes are best explained by transient transcriptional activation of fgr during TPA-induced differentiation, although nuclear runoff experiments were not sensitive enough to detect this event. Their results demonstrate that the c-fgr gene is expressed in a tissue- and development-specific fashion and suggest that constitutive expression of c-fgr in U937 cells is regulated by a labile transcriptional repressor.

  4. Tissue-Specific Regulation of Oncogene Expression Using Cre-Inducible ROSA26 Knock-In Transgenic Mice.

    PubMed

    Carofino, Brandi L; Justice, Monica J

    2015-01-01

    Cre-inducible mouse models are often utilized for the spatial and temporal expression of oncogenes. With the wide number of Cre recombinase lines available, inducible transgenesis represents a tractable approach to achieve discrete oncogene expression. Here, we describe a protocol for targeting Cre-inducible genes to the ubiquitously expressed ROSA26 locus. Gene targeting provides several advantages over standard transgenic techniques, including a known site of integration and previously characterized pattern of expression. Historically, an inherent instability of ROSA26 targeting vectors has hampered the efficiency of developing ROSA26 knock-in lines. In this protocol, we provide individual steps for utilizing Gateway recombination for cloning as well as detailed instructions for screening targeted ES cell clones. By following this protocol, one can achieve germline transmission of a ROSA26 knock-in line within several months. PMID:26069083

  5. Oncogenic KRAS Impairs EGFR Antibodies' Efficiency by C/EBPβ-Dependent Suppression of EGFR Expression12

    PubMed Central

    Derer, Stefanie; Berger, Sven; Schlaeth, Martin; Schneider-Merck, Tanja; Klausz, Katja; Lohse, Stefan; Overdijk, Marije B; Dechant, Michael; Kellner, Christian; Nagelmeier, Iris; Scheel, Andreas H; Lammerts van Bueren, Jeroen J; van de Winkel, Jan GJ; Parren, Paul WHI; Peipp, Matthias; Valerius, Thomas

    2012-01-01

    Oncogenic KRAS mutations in colorectal cancer (CRC) are associated with lack of benefit from epidermal growth factor receptor (EGFR)-directed antibody (Ab) therapy. However, the mechanisms by which constitutively activated KRAS (KRASG12V) impairs effector mechanisms of EGFR-Abs are incompletely understood. Here, we established isogenic cell line models to systematically investigate the impact of KRASG12V on tumor growth in mouse A431 xenograft models as well as on various modes of action triggered by EGFR-Abs in vitro. KRASG12V impaired EGFR-Ab-mediated growth inhibition by stimulating receptor-independent downstream signaling. KRASG12V also rendered tumor cells less responsive to Fc-mediated effector mechanisms of EGFR-Abs—such as complement-dependent cytotoxicity (CDC) and Ab-dependent cell-mediated cytotoxicity (ADCC). Impaired CDC and ADCC activities could be linked to reduced EGFR expression in KRAS-mutated versus wild-type (wt) cells, which was restored by small interfering RNA (siRNA)-mediated knockdown of KRAS4b. Immunohistochemistry experiments also revealed lower EGFR expression in KRAS-mutated versus KRAS-wt harboring CRC samples. Analyses of potential mechanisms by which KRASG12V downregulated EGFR expression demonstrated significantly decreased activity of six distinct transcription factors. Additional experiments suggested the CCAAT/enhancer-binding protein (C/EBP) family to be implicated in the regulation of EGFR promoter activity in KRAS-mutated tumor cells by suppressing EGFR transcription through up-regulation of the inhibitory family member C/EBPβ-LIP. Thus, siRNA-mediated knockdown of C/EBPβ led to enhanced EGFR expression and Ab-mediated cytotoxicity against KRAS-mutated cells. Together, these results demonstrate that KRASG12V signaling induced C/EBPβ-dependent suppression of EGFR expression, thereby impairing Fc-mediated effector mechanisms of EGFR-Abs and rendering KRAS-mutated tumor cells less sensitive to these therapeutic agents. PMID

  6. Regulatory mechanisms, expression levels and proliferation effects of the FUS-DDIT3 fusion oncogene in liposarcoma.

    PubMed

    Åman, Pierre; Dolatabadi, Soheila; Svec, David; Jonasson, Emma; Safavi, Setareh; Andersson, Daniel; Grundevik, Pernilla; Thomsen, Christer; Ståhlberg, Anders

    2016-04-01

    Fusion oncogenes are among the most common types of oncogene in human cancers. The gene rearrangements result in new combinations of regulatory elements and functional protein domains. Here we studied a subgroup of sarcomas and leukaemias characterized by the FET (FUS, EWSR1, TAF15) family of fusion oncogenes, including FUS-DDIT3 in myxoid liposarcoma (MLS). We investigated the regulatory mechanisms, expression levels and effects of FUS-DDIT3 in detail. FUS-DDIT3 showed a lower expression than normal FUS at both the mRNA and protein levels, and single-cell analysis revealed a lack of correlation between FUS-DDIT3 and FUS expression. FUS-DDIT3 transcription was regulated by the FUS promotor, while its mRNA stability depended on the DDIT3 sequence. FUS-DDIT3 protein stability was regulated by protein interactions through the FUS part, rather than the leucine zipper containing DDIT3 part. In addition, in vitro as well as in vivo FUS-DDIT3 protein expression data displayed highly variable expression levels between individual MLS cells. Combined mRNA and protein analyses at the single-cell level showed that FUS-DDIT3 protein expression was inversely correlated to the expression of cell proliferation-associated genes. We concluded that FUS-DDIT3 is uniquely regulated at the transcriptional as well as the post-translational level and that its expression level is important for MLS tumour development. The FET fusion oncogenes are potentially powerful drug targets and detailed knowledge about their regulation and functions may help in the development of novel treatments. PMID:26865464

  7. ZYG11A serves as an oncogene in non-small cell lung cancer and influences CCNE1 expression

    PubMed Central

    Wang, Xin; Sun, Qi; Chen, Chen; Yin, Rong; Huang, Xing; Wang, Xuan; Shi, Run; Xu, Lin; Ren, Binhui

    2016-01-01

    By analyzing The Cancer Genome Atlas (TCGA) database, we identified ZYG11A as a potential oncogene. We determined the expression of ZYG11A in NSCLC tissues and explored its clinical significance. And also evaluated the effects of ZYG11A on NSCLC cell proliferation, migration, and invasion both in vitro and in vivo. Our results show that ZYG11A is hyper-expressed in NSCLC tissues compared to adjacent normal tissues, and increased expression of ZYG11A is associated with a poor prognosis (HR: 2.489, 95%CI: 1.248-4.963, p = 0.010). ZYG11A knockdown induces cell cycle arrest and inhibits proliferation, migration, and invasion of NSCLC cells. ZYG11A knockdown also results in decreased expression of CCNE1. Over-expression of CCNE1 in cells with ZYG11A knockdown restores their oncogenic activities. Our data suggest that ZYG11A may serve as a novel oncogene promoting tumorigenicity of NSCLC cells by inducing cell cycle alterations and increasing CCNE1 expression. PMID:26771237

  8. The MYC 3′ Wnt-Responsive Element Drives Oncogenic MYC Expression in Human Colorectal Cancer Cells

    PubMed Central

    Rennoll, Sherri A.; Eshelman, Melanie A.; Raup-Konsavage, Wesley M.; Kawasawa, Yuka Imamura; Yochum, Gregory S.

    2016-01-01

    Mutations in components of the Wnt/β-catenin signaling pathway drive colorectal cancer (CRC) by deregulating expression of downstream target genes including the c-MYC proto-oncogene (MYC). The critical regulatory DNA enhancer elements that control oncogenic MYC expression in CRC have yet to be fully elucidated. In previous reports, we correlated T-cell factor (TCF) and β-catenin binding to the MYC 3′ Wnt responsive DNA element (MYC 3′ WRE) with MYC expression in HCT116 cells. Here we used CRISPR/Cas9 to determine whether this element is a critical driver of MYC. We isolated a clonal population of cells that contained a deletion of a single TCF binding element (TBE) within the MYC 3′ WRE. This deletion reduced TCF/β-catenin binding to this regulatory element and decreased MYC expression. Using RNA-Seq analysis, we found altered expression of genes that regulate metabolic processes, many of which are known MYC target genes. We found that 3′ WRE-Mut cells displayed a reduced proliferative capacity, diminished clonogenic growth, and a decreased potential to form tumors in vivo. These findings indicate that the MYC 3′ WRE is a critical driver of oncogenic MYC expression and suggest that this element may serve as a therapeutic target for CRC. PMID:27223305

  9. Merkel cell carcinoma subgroups by Merkel cell polyomavirus DNA relative abundance and oncogene expression

    PubMed Central

    Bhatia, Kishor; Goedert, James J.; Modali, Rama; Preiss, Liliana; Ayers, Leona W.

    2010-01-01

    Merkel cell polyomavirus (MCPyV) was recently discovered in Merkel cell carcinoma (MCC), a clinically and pathologically heterogeneous malignancy of dermal neuroendocrine cells. To investigate this heterogeneity, we developed a tissue microarray (TMA) to characterize immunohistochemical staining of candidate tumor cell proteins and a quantitative PCR assay to detect MCPyV and measure viral loads. MCPyV was detected in 19 of 23 (74%) primary MCC tumors, but 8 of these had less than 1 viral copy per 300 cells. Viral abundance of 0.06–1.2viral copies/cell was directly related to presence of retinoblastoma gene product (pRb) and terminal deoxyribonucleotidyl transferase (TdT) by immunohistochemical staining (P≤0.003). Higher viral abundance tumors tended to be associated with less p53 expression, younger age at diagnosis, and longer survival (P≤0.08). These data suggest that MCC may arise through different oncogenic pathways, including ones independent of pRb and MCPyV. PMID:19551862

  10. Immortality, but not oncogenic transformation, of primary human cells leads to epigenetic reprogramming of DNA methylation and gene expression

    PubMed Central

    Gordon, Katrina; Clouaire, Thomas; Bao, Xun X.; Kemp, Sadie E.; Xenophontos, Maria; de Las Heras, Jose Ignacio; Stancheva, Irina

    2014-01-01

    Tumourigenic transformation of normal cells into cancer typically involves several steps resulting in acquisition of unlimited growth potential, evasion of apoptosis and non-responsiveness to growth inhibitory signals. Both genetic and epigenetic changes can contribute to cancer development and progression. Given the vast genetic heterogeneity of human cancers and difficulty to monitor cancer-initiating events in vivo, the precise relationship between acquisition of genetic mutations and the temporal progression of epigenetic alterations in transformed cells is largely unclear. Here, we use an in vitro model system to investigate the contribution of cellular immortality and oncogenic transformation of primary human cells to epigenetic reprogramming of DNA methylation and gene expression. Our data demonstrate that extension of replicative life span of the cells is sufficient to induce accumulation of DNA methylation at gene promoters and large-scale changes in gene expression in a time-dependent manner. In contrast, continuous expression of cooperating oncogenes in immortalized cells, although essential for anchorage-independent growth and evasion of apoptosis, does not affect de novo DNA methylation at promoters and induces subtle expression changes. Taken together, these observations imply that cellular immortality promotes epigenetic adaptation to highly proliferative state, whereas transforming oncogenes confer additional properties to transformed human cells. PMID:24371281

  11. Expression of EBV-encoded oncogenes and EBV-like virions in multiple canine tumors.

    PubMed

    Chiu, Hung-Chuan; Chow, Kuan-Chih; Fan, Yi-Hsin; Chang, Shih-Chieh; Chiou, Shiow-Her; Chiang, Shu-Fen; Chiou, Che-Hao; Wu, Guo-Hua; Yang, Hsiu-Ching; Ho, Shu-Peng; Chen, Yuh-Kun; Lee, Wei-Cheng; Sun, H Sunny

    2013-04-12

    Epstein-Barr virus (EBV) is a ubiquitous human oncovirus. Previous studies by us and others have indicated that pet dogs frequently encounter EBV or EBV-related viral infection. In this study, we explored whether EBV is involved in canine malignancies in dogs. EBV-specific BamHI W sequence was detected by polymerase chain reaction (PCR) in 10 of 12 canine tumor specimens, including 8 of 10 oral tumors. Using reverse transcription-PCR, gene expressions of latent membrane protein 1 (LMP 1) and BamHI H rightward reading frame 1 (BHRF1) were identified in 8 and 7 of 12 specimens, respectively. A novel LMP1 variant, T0905, was predominant in 5 canine tumor specimens and found to exist in EBV positive human BC-2 cells. Another LMP1 variant, T0902, was similar to human tumor variant JB7. The BHRF1 sequence identified from these canine tumors was identical to that of the B95-8 viral strain. LMP1 protein and EBV-encoded RNA (EBER) were detected by immunohistochemistry and fluorescent in situ hybridization, respectively, in several tumors, particularly in tumor nests of oral amelanotic melanomas. Furthermore, EBV-like virions adopting a herpesvirus egress pathway were detected in a canthal fibroblastic osteosarcoma and an oral amelanotic melanoma. In conclusion, we report the expressions of BHRF1 transcript (a viral anti-apoptotic protein), LMP1 (a viral oncoprotein) transcript and protein, EBER (a viral oncogenic RNA), and EBV-like virions in multiple canine tumors. The identity of BHRF1 and the resemblance of LMP1 variants between canine and human tumors indicate either a close evolutionary relationship between canine and human EBV, or the possibility of zoonotic transmission. PMID:23380461

  12. Normal ABL1 is a tumor suppressor and therapeutic target in human and mouse leukemias expressing oncogenic ABL1 kinases.

    PubMed

    Dasgupta, Yashodhara; Koptyra, Mateusz; Hoser, Grazyna; Kantekure, Kanchan; Roy, Darshan; Gornicka, Barbara; Nieborowska-Skorska, Margaret; Bolton-Gillespie, Elisabeth; Cerny-Reiterer, Sabine; Müschen, Markus; Valent, Peter; Wasik, Mariusz A; Richardson, Christine; Hantschel, Oliver; van der Kuip, Heiko; Stoklosa, Tomasz; Skorski, Tomasz

    2016-04-28

    Leukemias expressing constitutively activated mutants of ABL1 tyrosine kinase (BCR-ABL1, TEL-ABL1, NUP214-ABL1) usually contain at least 1 normal ABL1 allele. Because oncogenic and normal ABL1 kinases may exert opposite effects on cell behavior, we examined the role of normal ABL1 in leukemias induced by oncogenic ABL1 kinases. BCR-ABL1-Abl1(-/-) cells generated highly aggressive chronic myeloid leukemia (CML)-blast phase-like disease in mice compared with less malignant CML-chronic phase-like disease from BCR-ABL1-Abl1(+/+) cells. Additionally, loss of ABL1 stimulated proliferation and expansion of BCR-ABL1 murine leukemia stem cells, arrested myeloid differentiation, inhibited genotoxic stress-induced apoptosis, and facilitated accumulation of chromosomal aberrations. Conversely, allosteric stimulation of ABL1 kinase activity enhanced the antileukemia effect of ABL1 tyrosine kinase inhibitors (imatinib and ponatinib) in human and murine leukemias expressing BCR-ABL1, TEL-ABL1, and NUP214-ABL1. Therefore, we postulate that normal ABL1 kinase behaves like a tumor suppressor and therapeutic target in leukemias expressing oncogenic forms of the kinase. PMID:26864341

  13. Expression of the c-myb proto-oncogene in bovine vascular smooth muscle cells.

    PubMed

    Brown, K E; Kindy, M S; Sonenshein, G E

    1992-03-01

    Previously we have shown that bovine vascular smooth muscle cells (SMCs) express c-myb mRNA (Reilly, C. F., Kindy, M. S., Brown, K. E., Rosenberg, R. D., and Sonenshein, G. E. (1989) J. Biol. Chem. 264, 6990-6995). Here we have characterized changes in the low level of c-myb mRNA expressed in quiescent serum-deprived subconfluent SMCs upon entry into the cell cycle. After serum stimulation, levels of c-myb mRNA increased 3-4-fold during late G1 and remained at this level during S phase. A 1.5-kilobase partial c-myb cDNA clone, isolated from a bovine SMC library, was partially sequenced and found to be 89 and 85% homologous to the human and murine c-myb genes, respectively. Using bovine and murine c-myb clones, no change in the rate of c-myb gene transcription or mRNA stability was detected during the cell cycle. Thus, the regulation of changes in c-myb mRNA levels in SMCs appears distinct from mechanisms seen in hematopoietic or fibroblastic cells. Vectors containing myb binding sites linked to the thymidine kinase promoter and the chloramphenicol acetyltransferase reporter gene were transiently transfected into SMC cultures. KHK-CAT-dAX, which contains nine concatenated myb binding sites, exhibited 7-fold more activity than the parental dAX-TK-CAT vector in exponentially growing SMCs. The levels of chloramphenicol acetyltransferase activity in exponentially growing cells were approximately 2-fold higher than in cells that had been serum deprived for 24 h and were entering quiescence. Thus SMCs produce a functional c-myb protein that can activate transcription from a heterologous promoter. Furthermore, introduction of antisense c-myb oligonucleotides to quiescent serum-deprived SMC cultures severely inhibited entry of cells into S phase upon serum addition. Thus, expression of the c-myb oncogene plays an important role in cell cycle progression of SMCs. PMID:1537845

  14. Transgenic expression of oncogenic BRAF induces loss of stem cells in the mouse intestine, which is antagonized by β-catenin activity.

    PubMed

    Riemer, P; Sreekumar, A; Reinke, S; Rad, R; Schäfer, R; Sers, C; Bläker, H; Herrmann, B G; Morkel, M

    2015-06-11

    Colon cancer cells frequently carry mutations that activate the β-catenin and mitogen-activated protein kinase (MAPK) signaling cascades. Yet how oncogenic alterations interact to control cellular hierarchies during tumor initiation and progression is largely unknown. We found that oncogenic BRAF modulates gene expression associated with cell differentiation in colon cancer cells. We therefore engineered a mouse with an inducible oncogenic BRAF transgene, and analyzed BRAF effects on cellular hierarchies in the intestinal epithelium in vivo and in primary organotypic culture. We demonstrate that transgenic expression of oncogenic BRAF in the mouse strongly activated MAPK signal transduction, resulted in the rapid development of generalized serrated dysplasia, but unexpectedly also induced depletion of the intestinal stem cell (ISC) pool. Histological and gene expression analyses indicate that ISCs collectively converted to short-lived progenitor cells after BRAF activation. As Wnt/β-catenin signals encourage ISC identity, we asked whether β-catenin activity could counteract oncogenic BRAF. Indeed, we found that intestinal organoids could be partially protected from deleterious oncogenic BRAF effects by Wnt3a or by small-molecule inhibition of GSK3β. Similarly, transgenic expression of stabilized β-catenin in addition to oncogenic BRAF partially prevented loss of stem cells in the mouse intestine. We also used BRAF(V637E) knock-in mice to follow changes in the stem cell pool during serrated tumor progression and found ISC marker expression reduced in serrated hyperplasia forming after BRAF activation, but intensified in progressive dysplastic foci characterized by additional mutations that activate the Wnt/β-catenin pathway. Our study suggests that oncogenic alterations activating the MAPK and Wnt/β-catenin pathways must be consecutively and coordinately selected to assure stem cell maintenance during colon cancer initiation and progression. Notably, loss of

  15. Regulation of proto-oncogene expression in adult and developing lungs.

    PubMed Central

    Molinar-Rode, R; Smeyne, R J; Curran, T; Morgan, J I

    1993-01-01

    Activation of immediate-early gene expression has been associated with mitogenesis, differentiation, nerve cell depolarization, and recently, terminal differentiation processes and programmed cell death. Previous evidence also suggested that immediate-early genes play a role in the physiology of the lungs (J. I. Morgan, D. R. Cohen, J. L. Hempstead, and T. Curran, Science 237:192-197, 1987). Therefore, we analyzed c-fos expression in adult and developing lung tissues. Seizures elicited by chemoconvulsants induced expression of mRNA for c-fos, c-jun, and junB and Fos-like immunoreactivity in lung tissue. The use of pharmacological antagonists and adrenalectomy indicated that this increased expression was neurogenic. Interestingly, by using a fos-lacZ transgenic mouse, it was shown that Fos-LacZ expression in response to seizure occurred preferentially in clusters of epithelial cells at the poles of the bronchioles. This was the same location of Fos-LacZ expression detected during early lung development. These data imply that pharmacological induction of immediate-early gene expression in adult mice recapitulates an embryological program of gene expression. Images PMID:8497249

  16. Neu proto-oncogene amplification and expression in ovarian adenocarcinoma cell lines.

    PubMed Central

    King, B. L.; Carter, D.; Foellmer, H. G.; Kacinski, B. M.

    1992-01-01

    In this communication, the authors summarize their characterization of eight ovarian adenocarcinoma-derived cell lines for level of neu gene amplification, expression of neu transcripts and protein, and intraperitoneal tumorigenicity in nude mice. Two of the eight cell lines in our study (SKOV3 and YAOVBIX1) exhibited five- to ninefold neu DNA sequence amplification, accompanied by up to 200-fold overexpression of transcripts and protein (p185). Both of these cell lines expressed a major approximately 7.5 kb neu-complementary transcript not previously reported in other neu-positive tumor cell lines. One pair of cell lines (YAOVBIX1 and YAOVBIX3), isolated from a single ovarian carcinoma patient's ascites sample differed dramatically in regard to level of neu gene amplification and expression. Immunohistochemical staining of the primary ovarian tumor from which these two lines were derived demonstrated populations of both neu-positive and neu-negative malignant epithelial cells. Seven of the eight ovarian carcinoma lines produced intra-abdominal tumors after intraperitoneal injection into nude mice, irrespective of level of neu gene expression. This study demonstrates tumor cell heterogeneity with regard to neu gene amplification and expression in an ovarian adenocarcinoma, reveals the overexpression of novel neu-complementary transcripts in two independently isolated ovarian adenocarcinoma cell lines, and suggests that neu gene expression is not required for intraperitoneal tumorigenicity of ovarian carcinoma xenografts in a nude mouse model system. Images Figure 4 Figure 1 Figure 2 Figure 3 PMID:1346236

  17. Synergistic Induction of Potential Warburg Effect in Zebrafish Hepatocellular Carcinoma by Co-Transgenic Expression of Myc and xmrk Oncogenes

    PubMed Central

    Li, Zhen; Zheng, Weiling; Li, Hankun; Li, Caixia; Gong, Zhiyuan

    2015-01-01

    Previously we have generated inducible liver tumor models by transgenic expression of Myc or xmrk (activated EGFR homolog) oncogenes in zebrafish. To investigate the interaction of the two oncogenes, we crossed the two transgenic lines and observed more severe and faster hepatocarcinogenesis in Myc/xmrk double transgenic zebrafish than either single transgenic fish. RNA-Seq analyses revealed distinct changes in many molecular pathways among the three types of liver tumors. In particular, we found dramatic alteration of cancer metabolism based on the uniquely enriched pathways in the Myc/xmrk tumors. Critical glycolytic genes including hk2, pkm and ldha were significantly up-regulated in Myc/xmrk tumors but not in either single oncogene-induced tumors, suggesting a potential Warburg effect. In RT-qPCR analyses, the specific pkm2 isoformin Warburg effect was found to be highly enriched in the Myc/xmrk tumors but not in Myc or xmrk tumors, consistent with the observations in many human cancers with Warburg effect. Moreover, the splicing factor genes (hnrnpa1, ptbp1a, ptbp1b and sfrs3b) responsible for generating the pkm isoform were also greatly up-regulated in the Myc/xmrk tumors. As Pkm2 isoform is generally inactive and causes incomplete glycolysis to favor anabolism and tumor growth, by treatment with a Pkm2-specific activator, TEPP-46, we further demonstrated that activation of Pkm2 suppressed the growth of oncogenic liver as well as proliferation of liver cells. Collectively, our Myc/xmrk zebrafish model suggests synergetic effect of EGFR and MYC in triggering Warburg effect in the HCC formation and may provide a promising in vivo model for Warburg effect. PMID:26147004

  18. Suppression of c-myc oncogene expression by a polyamine-complexed triplex forming oligonucleotide in MCF-7 breast cancer cells.

    PubMed Central

    Thomas, T J; Faaland, C A; Gallo, M A; Thomas, T

    1995-01-01

    Polyamines are excellent stabilizers of triplex DNA. Recent studies in our laboratory revealed a remarkable structural specificity of polyamines in the induction and stabilization of triplex DNA. 1,3-Diaminopropane (DAP) showed optimum efficacy amongst a series of synthetic diamines in stabilizing triplex DNA. To utilize the potential of this finding in developing an anti-gene strategy for breast cancer, we treated MCF-7 cells with a 37mer oligonucleotide to form triplex DNA in the up-stream regulatory region of the c-myc oncogene in the presence of DAP. As individual agents, the oligonucleotide and DAP did not downregulate c-myc mRNA in the presence of estradiol. Complexation of the oligonucleotide with 2 mM DAP reduced c-myc mRNA signal by 65% at 10 microM oligonucleotide concentration. In contrast, a control oligonucleotide had no significant effect on c-myc mRNA. The expression of c-fos oncogene was not significantly altered by the triplex forming oligonucleotide (TFO). DAP was internalized within 1 h of treatment; however, it had no significant effect on the level of natural polyamines. These data indicate that selective utilization of synthetic polyamines and TFOs might be an important strategy to develop anti-gene-based therapeutic modalities for breast cancer. Images PMID:7567474

  19. Immunohistochemichal Assessment of the CrkII Proto-oncogene Expression in Common Malignant Salivary Gland Tumors and Pleomorphic Adenoma.

    PubMed

    Askari, Mitra; Darabi, Masoud; Jahanzad, Esa; Mostakhdemian Hosseini, Zahra; Musavi Chavoshi, Marjan; Darabi, Maryam

    2015-01-01

    Background and aims. Various morphologies are seen in different salivary gland tumorsor within an individual tumor, and the lesions show divers biological behaviors. Experimental results support the hypothesis that increased CrkII proto-oncogene is associated with cytokine-induced tumor initiation and progression by altering cell motility signaling pathway. The aim of this study was to assess the CrkII expression in common malignant salivary gland tumors and pleomorphic ade-noma. Materials and methods. Immunohistochemical analysis of CrkII expression was performed on paraffin blocks of 64 car-cinomas of salivary glands, 10 pleomorphic adenomas, and 10 normal salivary glands. Biopsies were subjected to immu-nostaining with EnVision detection system using monoclonal anti-CrkII. Evaluation of immunoreactivity of CrkII was based on the immunoreaction intensity and percentage of stained tumor cells which were scored semi-quantitatively on a scale with four grades 0 to 3. Kruskal-wallis test and additional Mann-Whitney statistical test were used for analysis of CrkII expression levels. Results. Increased expression of CrkII was seen (P=0.005) in malignant tumors including: mucoepidermoid carcinoma, adenoid cystic carcinoma, and carcinoma ex pleomorphic adenoma, but CrkII expression in acinic cell carcinoma was weak. CrkII expression in pleomorphic adenoma was weak or negative. A weak staining was sparsely seen in normal acinar serous cell. Conclusion. Increased expression of CrkII and its higher intensity of staining in tumors with more aggressive biologic behavior in carcinomas of salivary gland is consistent with a role for this proto-oncogene in salivary gland tumorigenesis and cancer progression. PMID:25973151

  20. Initiation of transcription from the minute virus of mice P4 promoter is stimulated in rat cells expressing a c-Ha-ras oncogene.

    PubMed Central

    Spegelaere, P; van Hille, B; Spruyt, N; Faisst, S; Cornelis, J J; Rommelaere, J

    1991-01-01

    Transformation of FR3T3 rat fibroblasts by a c-Ha-ras oncogene but not by bovine papillomavirus type 1 is associated with an increase in the abundance of mRNAs from prototype strain MVMp of infecting minute virus of mice, an oncosuppressive parvovirus. This differential parvovirus gene expression correlates with the reported sensitization of ras- but not bovine papillomavirus type 1-transformed cells to the killing effect of MVMp (N. Salomé, B. van Hille, N. Duponchel, G. Meneguzzi, F. Cuzin, J. Rommelaere, and J. Cornelis, Oncogene 5:123-130, 1990). Experiments were performed to determine at which level parvovirus expression is up-regulated in ras transformants. An MVMp "attenuation" sequence responsible for the premature arrest of RNA elongation was either placed or not placed in front of the chloramphenicol acetyltransferase gene and brought under the control of MVMp early promoter P4. Although the MVMp attenuator reduced P4-driven chloramphenicol acetyltransferase expression, the extent of attenuation was similar in normal and ras-transformed cells. Moreover, the analysis of P4-directed viral RNAs in MVMp-infected cultures by RNase protection and nuclear run-on assays also revealed a transcription elongation block of a similar amplitude in both types of cells. In addition, the stabilities of the three major parvoviral mRNAs did not vary significantly between normal and ras-transformed cells. Hence, it is concluded that the ras-induced increase in the accumulation of parvoviral mRNAs is mainly controlled at the level of transcription. Consistently, the TATA motif of the P4 promoter proved to have a differential photoreactivity when tested by in vivo UV footprinting assays in ras-transformed versus normal cells. Images PMID:1651412

  1. The expression of heat shock protein hsp27 and a complexed 22-kilodalton protein is inversely correlated with oncogenicity of adenovirus-transformed cells.

    PubMed Central

    Zantema, A; de Jong, E; Lardenoije, R; van der Eb, A J

    1989-01-01

    We isolated a monoclonal antibody that immunoprecipitated two proteins of 22 and 27 kilodaltons (kDa) from nononcogenic adenovirus type 5 early region 1 (E1)-transformed rat cells but not from oncogenic adenovirus type 12 E1-transformed rat cells. In a variety of adenovirus-transformed cells including cells transformed by E1A and the c-H-ras oncogene, we found a perfect, inverse correlation between the presence of these two proteins and the oncogenicity of these cells in syngeneic immunocompetent rats. Characterization of the two proteins revealed that they occur in a large (700-kDa) complex and that the 27-kDa protein is identical to the already known 27-kDa (28-kDa) heat shock protein hsp27. The suppression of the hsp27 protein in oncogenic cells is further demonstrated by the fact that its mRNA is absent even after heat-shock induction. Images PMID:2746733

  2. Expression of the human ETS-2 oncogene in normal fetal tissues and in the brain of a fetus with trisomy 21.

    PubMed

    Baffico, M; Perroni, L; Rasore-Quartino, A; Scartezzini, P

    1989-10-01

    The expression of the ETS-2 proto-oncogene, located on chromosome 21, in normal fetal tissues and in neural tissue of a fetus affected by Down syndrome has been investigated. The results show that the ETS-2 proto-oncogene is expressed in almost all the tissues examined and that it is transcribed at constant levels in neural tissue between the 13th and 24th weeks. ETS-2 expression appeared to be slightly increased in Down syndrome brain compared with that of normal controls of the same gestational age. PMID:2529204

  3. mTORC1 upregulation via ERK-dependent gene expression change confers intrinsic resistance to MEK inhibitors in oncogenic KRas-mutant cancer cells.

    PubMed

    Komatsu, N; Fujita, Y; Matsuda, M; Aoki, K

    2015-11-01

    Cancer cells harboring oncogenic BRaf mutants, but not oncogenic KRas mutants, are sensitive to MEK inhibitors (MEKi). The mechanism underlying the intrinsic resistance to MEKi in KRas-mutant cells is under intensive investigation. Here, we pursued this mechanism by live imaging of extracellular signal-regulated kinases (ERK) and mammalian target of rapamycin complex 1 (mTORC1) activities in oncogenic KRas or BRaf-mutant cancer cells. We established eight cancer cell lines expressing Förster resonance energy transfer (FRET) biosensors for ERK activity and S6K activity, which was used as a surrogate marker for mTORC1 activity. Under increasing concentrations of MEKi, ERK activity correlated linearly with the cell growth rate in BRaf-mutant cancer cells, but not KRas-mutant cancer cells. The administration of PI3K inhibitors resulted in a linear correlation between ERK activity and cell growth rate in KRas-mutant cancer cells. Intriguingly, mTORC1 activity was correlated linearly with the cell growth rate in both BRaf-mutant cancer cells and KRas-mutant cancer cells. These observations suggested that mTORC1 activity had a pivotal role in cell growth and that the mTORC1 activity was maintained primarily by the ERK pathway in BRaf-mutant cancer cells and by both the ERK and PI3K pathways in KRas-mutant cancer cells. FRET imaging revealed that MEKi inhibited mTORC1 activity with slow kinetics, implying transcriptional control of mTORC1 activity by ERK. In agreement with this observation, MEKi induced the expression of negative regulators of mTORC1, including TSC1, TSC2 and Deptor, which occurred more significantly in BRaf-mutant cells than in KRas-mutant cells. These findings suggested that the suppression of mTORC1 activity and induction of negative regulators of mTORC1 in cancer cells treated for at least 1 day could be used as surrogate markers for the MEKi sensitivity of cancer cells. PMID:25703330

  4. Flavopiridol induces BCL-2 expression and represses oncogenic transcription factors in leukemic blasts from adults with refractory acute myeloid leukemia

    PubMed Central

    Nelson, Dwella M.; Joseph, Biju; Hillion, Joelle; Segal, Jodi; Karp, Judith E.; Resar, Linda M. S.

    2011-01-01

    Flavopiridol is a cyclin-dependent kinase inhibitor that induces cell cycle arrest, apoptosis, and clinical responses in selected patients with acute myeloid leukemia (AML). A better understanding of the molecular pathways targeted by flavopiridol is needed to design optimal combinatorial therapy. Here, we report that in vivo administration of flavopiridol induced expression of the BCL-2 anti-apoptotic gene in leukemic blasts from adult patients with refractory AML. Moreover, flavopiridol repressed the expression of genes encoding oncogenic transcription factors (HMGA1, STAT3, E2F1) and the major subunit of RNA Polymerase II. Our results provide mechanistic insight into the cellular pathways targeted by flavopiridol and suggest that blocking anti-apoptotic pathways could enhance cytotoxicity and improve outcomes in patients treated with flavopiridol. PMID:21728742

  5. Cell-cycle dependent expression of a translocation-mediated fusion oncogene mediates checkpoint adaptation in rhabdomyosarcoma.

    PubMed

    Kikuchi, Ken; Hettmer, Simone; Aslam, M Imran; Michalek, Joel E; Laub, Wolfram; Wilky, Breelyn A; Loeb, David M; Rubin, Brian P; Wagers, Amy J; Keller, Charles

    2014-01-01

    Rhabdomyosarcoma is the most commonly occurring soft-tissue sarcoma in childhood. Most rhabdomyosarcoma falls into one of two biologically distinct subgroups represented by alveolar or embryonal histology. The alveolar subtype harbors a translocation-mediated PAX3:FOXO1A fusion gene and has an extremely poor prognosis. However, tumor cells have heterogeneous expression for the fusion gene. Using a conditional genetic mouse model as well as human tumor cell lines, we show that that Pax3:Foxo1a expression is enriched in G2 and triggers a transcriptional program conducive to checkpoint adaptation under stress conditions such as irradiation in vitro and in vivo. Pax3:Foxo1a also tolerizes tumor cells to clinically-established chemotherapy agents and emerging molecularly-targeted agents. Thus, the surprisingly dynamic regulation of the Pax3:Foxo1a locus is a paradigm that has important implications for the way in which oncogenes are modeled in cancer cells. PMID:24453992

  6. The Circular RNA Cdr1as Act as an Oncogene in Hepatocellular Carcinoma through Targeting miR-7 Expression.

    PubMed

    Yu, Lei; Gong, Xuejun; Sun, Lei; Zhou, Qiying; Lu, Baoling; Zhu, Liying

    2016-01-01

    CircRNAs are a class of endogenous RNA that regulates gene expression at the post-transcriptional or transcriptionallevel through interacting with other molecules or microRNAs. Increasing studies have demonstrated that circRNAs play a crucial role in biology processes. CircRNAs are proved as potentialbiomarkers in many diseases including cancers. However, the role of Cdr1as in Hepatocellular carcinoma (HCC) remains to be elucidated. We demonstrated that Cdr1as expression was upregulated in HCC tissues compared with the adjacent non-tumor tissues. In addtion, miR-7 expression was downregulated in HCC tissues compared with the adjacent non-tumor tissues. Moreover, the expression level of miR-7 was inversely correlated with that in HCC tissues. Knockdown of Cdr1as suppressed the HCC cell proliferation and invasion. Overexpression of miR-7 inhibited the HCC cell proliferation and invasion. Overexpression of miR-7 could suppress the direct target gene CCNE1 and PIK3CD expression. Knockdown of Cdr1as suppressed the expression of miR-7 and also inhibited the CCNE1 and PIK3CD expression. Furthermore, knockdown of Cdr1as suppressed the HCC cell proliferation and invasion through targeting miR-7. These data suggested that Cdr1as acted as an oncogene partly through targeting miR-7 in HCC. PMID:27391479

  7. The Circular RNA Cdr1as Act as an Oncogene in Hepatocellular Carcinoma through Targeting miR-7 Expression

    PubMed Central

    Yu, Lei; Gong, Xuejun; Sun, Lei; Zhou, Qiying; Lu, Baoling; Zhu, Liying

    2016-01-01

    CircRNAs are a class of endogenous RNA that regulates gene expression at the post-transcriptional or transcriptionallevel through interacting with other molecules or microRNAs. Increasing studies have demonstrated that circRNAs play a crucial role in biology processes. CircRNAs are proved as potentialbiomarkers in many diseases including cancers. However, the role of Cdr1as in Hepatocellular carcinoma (HCC) remains to be elucidated. We demonstrated that Cdr1as expression was upregulated in HCC tissues compared with the adjacent non-tumor tissues. In addtion, miR-7 expression was downregulated in HCC tissues compared with the adjacent non-tumor tissues. Moreover, the expression level of miR-7 was inversely correlated with that in HCC tissues. Knockdown of Cdr1as suppressed the HCC cell proliferation and invasion. Overexpression of miR-7 inhibited the HCC cell proliferation and invasion. Overexpression of miR-7 could suppress the direct target gene CCNE1 and PIK3CD expression. Knockdown of Cdr1as suppressed the expression of miR-7 and also inhibited the CCNE1 and PIK3CD expression. Furthermore, knockdown of Cdr1as suppressed the HCC cell proliferation and invasion through targeting miR-7. These data suggested that Cdr1as acted as an oncogene partly through targeting miR-7 in HCC. PMID:27391479

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

    PubMed Central

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

    2001-01-01

    Background 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. 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. PMID:11231886

  9. Sterol Regulatory Element Binding Protein Regulates the Expression and Metabolic Functions of Wild-Type and Oncogenic IDH1.

    PubMed

    Ricoult, Stéphane J H; Dibble, Christian C; Asara, John M; Manning, Brendan D

    2016-09-15

    Sterol regulatory element binding protein (SREBP) is a major transcriptional regulator of the enzymes underlying de novo lipid synthesis. However, little is known about the SREBP-mediated control of processes that indirectly support lipogenesis, for instance, by supplying reducing power in the form of NAPDH or directing carbon flux into lipid precursors. Here, we characterize isocitrate dehydrogenase 1 (IDH1) as a transcriptional target of SREBP across a spectrum of cancer cell lines and human cancers. IDH1 promotes the synthesis of lipids specifically from glutamine-derived carbons. Neomorphic mutations in IDH1 occur frequently in certain cancers, leading to the production of the oncometabolite 2-hydroxyglutarate (2-HG). We found that SREBP induces the expression of oncogenic IDH1 and influences 2-HG production from glucose. Treatment of cells with 25-hydroxycholesterol or statins, which respectively inhibit or activate SREBP, further supports SREBP-mediated regulation of IDH1 and, in cells with oncogenic IDH1, carbon flux into 2-HG. PMID:27354064

  10. Oncogenic K-ras confers SAHA resistance by up-regulating HDAC6 and c-myc expression

    PubMed Central

    Zhang, Yi; Tan, Zhiping; Su, Bing; Li, Yu

    2016-01-01

    Histone deacetylase inhibitors (HDIs) represent a new class of anticancer drugs. Suberoylanilide hydroxamic acid (SAHA), the first HDI approved for the treatment of cutaneous T cell lymphoma (CTCL), is currently being tested in clinical trials for other cancers. However, SAHA has been ineffective against solid tumors in many clinical trials. A better understanding of molecular mechanisms of SAHA resistance may provide the basis for improved patient selection and the enhancement of clinical efficacy. Here we demonstrate that oncogenic K-ras contributes to SAHA resistance by upregulating HDAC6 and c-myc expression. We find that the high levels of HDAC6 expression are associated with activated K-ras mutant in colon cancer patients. And expressions of HDAC6 and c-myc are increased in fibroblasts transformed with activated K-ras. Surprisingly, we find that activated K-ras transformed cells are more resistant to SAHA inhibition on cell growth and anchorage-independent colony formation. We show that a K-ras inhibitor sensitizes K-ras mutated lung cancer cells to SAHA induced growth inhibition. We also find that mutant K-ras induces HDAC6 expression by a MAP kinase dependent pathway. Our study suggests that combined treatment with SAHA and K-ras inhibitors may represent an effective strategy to overcome SAHA resistance. PMID:26848526

  11. Oncogenic Activity of miR-650 in Prostate Cancer Is Mediated by Suppression of CSR1 Expression

    PubMed Central

    Zuo, Ze-Hua; Yu, Yan P.; Ding, Ying; Liu, Silvia; Martin, Amantha; Tseng, George; Luo, Jian-Hua

    2016-01-01

    Cellular stress response 1 (CSR1) is a tumor suppressor gene whose expression was frequently down-regulated in prostate cancer. The mechanism of its down-regulation, however, is not clear. Here, we show that the 3′ untranslated region of CSR1 contains a target site of miR-650. High level of miR-650 was found in prostate cancer samples and cell lines. Degradation of miR-650 by specific inhibitor dramatically increased the expression levels of CSR1. Interaction between miR-650 and its target site in the 3′ untranslated region was validated through luciferase reporter system. Mutation at the target site completely abrogated the activity of miR-650 on the 3′ untranslated region of CSR1. Inhibition of miR-650 reversed the expression suppression of CSR1, suppressed colony formation, and blocked cell cycle entry to the S phase of both PC3 and DU145 cells. Animal model showed significant decrease of tumor volume, rate of metastasis, and mortality of severe combined immunodeficient mice xenografted with PC3 or DU145 cells transformed with inhibitor of miR-650. Our analyses demonstrate that suppression of CSR1 expression is a novel mechanism critical for the oncogenic activity of miR-650. PMID:25956032

  12. Oncogenic Activity of miR-650 in Prostate Cancer Is Mediated by Suppression of CSR1 Expression.

    PubMed

    Zuo, Ze-Hua; Yu, Yan P; Ding, Ying; Liu, Silvia; Martin, Amantha; Tseng, George; Luo, Jian-Hua

    2015-07-01

    Cellular stress response 1 (CSR1) is a tumor suppressor gene whose expression was frequently down-regulated in prostate cancer. The mechanism of its down-regulation, however, is not clear. Here, we show that the 3' untranslated region of CSR1 contains a target site of miR-650. High level of miR-650 was found in prostate cancer samples and cell lines. Degradation of miR-650 by specific inhibitor dramatically increased the expression levels of CSR1. Interaction between miR-650 and its target site in the 3' untranslated region was validated through luciferase reporter system. Mutation at the target site completely abrogated the activity of miR-650 on the 3' untranslated region of CSR1. Inhibition of miR-650 reversed the expression suppression of CSR1, suppressed colony formation, and blocked cell cycle entry to the S phase of both PC3 and DU145 cells. Animal model showed significant decrease of tumor volume, rate of metastasis, and mortality of severe combined immunodeficient mice xenografted with PC3 or DU145 cells transformed with inhibitor of miR-650. Our analyses demonstrate that suppression of CSR1 expression is a novel mechanism critical for the oncogenic activity of miR-650. PMID:25956032

  13. Oncogenic K-ras confers SAHA resistance by up-regulating HDAC6 and c-myc expression.

    PubMed

    Wang, Qun; Tan, Rong; Zhu, Xin; Zhang, Yi; Tan, Zhiping; Su, Bing; Li, Yu

    2016-03-01

    Histone deacetylase inhibitors (HDIs) represent a new class of anticancer drugs. Suberoylanilide hydroxamic acid (SAHA), the first HDI approved for the treatment of cutaneous T cell lymphoma (CTCL), is currently being tested in clinical trials for other cancers. However, SAHA has been ineffective against solid tumors in many clinical trials. A better understanding of molecular mechanisms of SAHA resistance may provide the basis for improved patient selection and the enhancement of clinical efficacy. Here we demonstrate that oncogenic K-ras contributes to SAHA resistance by upregulating HDAC6 and c-myc expression. We find that the high levels of HDAC6 expression are associated with activated K-ras mutant in colon cancer patients. And expressions of HDAC6 and c-myc are increased in fibroblasts transformed with activated K-ras. Surprisingly, we find that activated K-ras transformed cells are more resistant to SAHA inhibition on cell growth and anchorage-independent colony formation. We show that a K-ras inhibitor sensitizes K-ras mutated lung cancer cells to SAHA induced growth inhibition. We also find that mutant K-ras induces HDAC6 expression by a MAP kinase dependent pathway. Our study suggests that combined treatment with SAHA and K-ras inhibitors may represent an effective strategy to overcome SAHA resistance. PMID:26848526

  14. Long Noncoding RNA MALAT1 Controls Cell Cycle Progression by Regulating the Expression of Oncogenic Transcription Factor B-MYB

    PubMed Central

    Tripathi, Vidisha; Shen, Zhen; Chakraborty, Arindam; Giri, Sumanprava; Freier, Susan M.; Wu, Xiaolin; Zhang, Yongqing; Gorospe, Myriam; Prasanth, Supriya G.; Lal, Ashish; Prasanth, Kannanganattu V.

    2013-01-01

    The long noncoding MALAT1 RNA is upregulated in cancer tissues and its elevated expression is associated with hyper-proliferation, but the underlying mechanism is poorly understood. We demonstrate that MALAT1 levels are regulated during normal cell cycle progression. Genome-wide transcriptome analyses in normal human diploid fibroblasts reveal that MALAT1 modulates the expression of cell cycle genes and is required for G1/S and mitotic progression. Depletion of MALAT1 leads to activation of p53 and its target genes. The cell cycle defects observed in MALAT1-depleted cells are sensitive to p53 levels, indicating that p53 is a major downstream mediator of MALAT1 activity. Furthermore, MALAT1-depleted cells display reduced expression of B-MYB (Mybl2), an oncogenic transcription factor involved in G2/M progression, due to altered binding of splicing factors on B-MYB pre-mRNA and aberrant alternative splicing. In human cells, MALAT1 promotes cellular proliferation by modulating the expression and/or pre-mRNA processing of cell cycle–regulated transcription factors. These findings provide mechanistic insights on the role of MALAT1 in regulating cellular proliferation. PMID:23555285

  15. Expression of the c-myc oncogene under control of an immunoglobulin enhancer in E mu-myc transgenic mice.

    PubMed

    Alexander, W S; Schrader, J W; Adams, J M

    1987-04-01

    Transgenic mice bearing a cellular myc oncogene coupled to the immunoglobulin heavy-chain enhancer (E mu) exhibit perturbed B-lymphocyte development and succumb to B lymphoid tumors. To investigate how the enhancer has affected myc expression, we analyzed the structure and abundance of myc transcripts in tissues of prelymphomatous mice and in the lymphomas. Expression of the E mu-myc transgene appeared to be confined largely to B lymphoid cells, being dominant in bone marrow, spleen, and lymph nodes, with no detectable expression in T cells or other hematopoietic lineages examined. The myc transcripts initiated very predominantly at the normal myc promoters, although use of the more upstream myc promoter was accentuated and an enhancer-associated promoter may be used infrequently. The level of E mu-myc transcripts in the preneoplastic lymphoid tissues and in the E mu-myc tumors was not markedly higher than myc RNA levels in proliferating normal lymphocytes. Thus, enforced expression of structurally normal myc transcripts at only a modestly elevated level has profound biological consequences. The absence of detectable endogenous c-myc RNA in any tumor, or in preneoplastic bone marrow, supports a negative feedback model for normal c-myc regulation. PMID:3037318

  16. Long noncoding RNA XIST acts as an oncogene in non-small cell lung cancer by epigenetically repressing KLF2 expression.

    PubMed

    Fang, Jing; Sun, Cheng-Cao; Gong, Cheng

    2016-09-16

    Recently, long noncoding RNAs (lncRNAs) have been identified as critical regulators in numerous types of cancers, including non-small cell lung cancer (NSCLC). X inactivate-specific transcript (XIST) has been found to be up-regulated and acts as an oncogene in gastric cancer and hepatocellular carcinoma, but little is known about its expression pattern, biological function and underlying mechanism in NSCLC. Here, we identified XIST as an oncogenic lncRNA by driving tumorigenesis in NSCLC. We found that XIST is over-expressed in NSCLC, and its increased level is associated with shorter survival and poorer prognosis. Knockdown of XIST impaired NSCLC cells proliferation, migration and invasion in vitro, and repressed the tumorigenicity of NSCLC cells in vivo. Mechanistically, RNA immune-precipitation (RIP) and RNA pull-down experiment demonstrated that XIST could simultaneously interact with EZH2 to suppress transcription of its potential target KLF2. Additionally, rescue experiments revealed that XIST's oncogenic functions were partly depending on silencing KLF2 expression. Collectively, our findings expound how XIST over-expression endows an oncogenic function in NSCLC. PMID:27501756

  17. Transient expression of Bcl6 is sufficient for oncogenic function and induction of mature B-cell lymphoma

    PubMed Central

    Green, Michael R; Vicente-Dueñas, Carolina; Romero-Camarero, Isabel; Liu, Chih Long; Dai, Bo; González-Herrero, Inés; García-Ramírez, Idoia; Alonso-Escudero, Esther; Iqbal, Javeed; Chan, Wing C; Campos-Sanchez, Elena; Orfao, Alberto; Pintado, Belén; Flores, Teresa; Blanco, Oscar; Jiménez, Rafael; Martínez-Climent, Jose Angel; Criado, Francisco Javier García; Cenador, María Begoña García; Zhao, Shuchun; Natkunam, Yasodha; Lossos, Izidore S; Majeti, Ravindra; Melnick, Ari; Cobaleda, César; Alizadeh, Ash A.; Sánchez-García, Isidro

    2015-01-01

    Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoma and can be separated into two subtypes based upon molecular features with similarities to germinal center B-cells (GCB-like) or activated B-cells (ABC-like). Here we identify gain of 3q27.2 as being significantly associated with adverse outcome in DLBCL and linked with the ABC-like subtype. This lesion includes the BCL6 oncogene, but does not alter BCL6 transcript levels or target-gene repression. Separately, we identify expression of BCL6 in a subset of human hematopoietic stem/progenitor cells (HSPCs). We therefore hypothesize that BCL6 may act by hit-and-run oncogenesis. We model this by transiently expressing Bcl6 within murine HSPCs, and find it causes mature B-cell lymphomas that lack Bcl6 expression and target-gene repression, are transcriptionally similar to post-GCB cells, and show epigenetic changes that are conserved from HSPCs to mature B-cells. Together these results suggest that Bcl6 may function in a hit-and-run role in lymphomagenesis. PMID:24887457

  18. Current Protocols in Mouse Biology Tissue-specific regulation of oncogene expression using Cre-inducible ROSA26 knock-in transgenic mice

    PubMed Central

    Carofino, Brandi L.; Justice, Monica J.

    2015-01-01

    Cre-inducible mouse models are often utilized for the spatial and temporal expression of oncogenes. With the wide number of Cre recombinase lines available, inducible transgenesis represents a tractable approach to achieve discrete oncogene expression. Here, we describe a protocol for targeting Cre-inducible genes using a loxP-STOP-loxP approach to the ubiquitously expressed ROSA26 locus. Gene targeting provides several advantages over standard transgenic techniques, including a known site of integration and previously characterized pattern of expression. Historically, an inherent instability of ROSA26 targeting vectors has hampered the efficiency of developing ROSA26 knock-in lines. In this protocol, we provide individual steps for utilizing Gateway recombination for cloning, and detailed instructions for screening targeted ES cell clones. By following this protocol, one can achieve germline transmission of a ROSA26 knock-in line within several months. PMID:26069083

  19. Expression of galaxin and oncogene homologs in growth anomaly in the coral Montipora capitata.

    PubMed

    Spies, Narrissa P; Takabayashi, Misaki

    2013-06-13

    Growth anomaly (GA) is a coral disease characterized by enlarged skeletal lesions. Although negative effects of GA on several of coral's biological functions have been determined, the etiology and molecular pathology of this disease is very poorly understood. We studied the expression of 5 genes suspected to play a role in pathological development of GA in the endemic Hawaiian coral Montipora capitata, which is particularly susceptible to this disease. Transcript abundances of the 5 target genes in healthy tissue, GA-affected tissue, and unaffected tissue (apparently healthy tissue adjacent to GA) relative to 3 internal control genes (actin, NADH, and rpS3) were compared using quantitative reverse transcriptase PCR. Galaxin, which codes for a protein suspected to be involved in calcification and thus hypothesized to be differentially expressed in GA, was up-regulated in unaffected tissue but remained at baseline levels in GA tissue. The gene expressions of murine double minute 2 (MDM2) and tumor necrosis factor (TNF) remained unchanged in GA tissue. The expression of tyrosine protein kinase (TPK) and βγ-crystallin (BGC) were both down-regulated. These expression patterns were all inconsistent with the expression patterns of homologous genes in neoplastic diseases featuring similar morphological symptoms in humans. These expression data therefore suggest that the calcification mechanism is likely not enhanced in coral GA and that coral GA is not a malignant neoplasia. PMID:23759562

  20. Oncogenes and growth control

    SciTech Connect

    Kahn, P.; Graf, T.

    1986-01-01

    This book contains six sections, each consisting of several papers. Some of the paper titles are: A Role for Proto-Oncogenes in Differentiation.; The ras Gene Family; Regulation of Human Globin Gene Expression; Regulation of Gene Expression by Steroid Hormones; The Effect of DNA Methylation on DNA-Protein Interactions and on the Regulation of Gene Expression; and Trans-Acting Elements Encoded in Immediate Early Genes of DNA Tumor Viruses.

  1. Expression of MALT1 oncogene in hematopoietic stem/progenitor cells recapitulates the pathogenesis of human lymphoma in mice

    PubMed Central

    Vicente-Dueñas, Carolina; Fontán, Lorena; Gonzalez-Herrero, Ines; Romero-Camarero, Isabel; Segura, Victor; Aznar, M. Angela; Alonso-Escudero, Esther; Campos-Sanchez, Elena; Ruiz-Roca, Lucía; Barajas-Diego, Marcos; Sagardoy, Ainara; Martinez-Ferrandis, Jose I.; Abollo-Jimenez, Fernando; Bertolo, Cristina; Peñuelas, Ivan; Garcia-Criado, Francisco J.; García-Cenador, María B.; Tousseyn, Thomas; Agirre, Xabier; Prosper, Felipe; Garcia-Bragado, Federico; McPhail, Ellen D.; Lossos, Izidore S.; Du, Ming-Qing; Flores, Teresa; Hernandez-Rivas, Jesus M.; Gonzalez, Marcos; Salar, Antonio; Bellosillo, Beatriz; Conde, Eulogio; Siebert, Reiner; Sagaert, Xavier; Cobaleda, Cesar; Sanchez-Garcia, Isidro; Martinez-Climent, Jose A.

    2012-01-01

    Chromosomal translocations involving the MALT1 gene are hallmarks of mucosa-associated lymphoid tissue (MALT) lymphoma. To date, targeting these translocations to mouse B cells has failed to reproduce human disease. Here, we induced MALT1 expression in mouse Sca1+Lin− hematopoietic stem/progenitor cells, which showed NF-κB activation and early lymphoid priming, being selectively skewed toward B-cell differentiation. These cells accumulated in extranodal tissues and gave rise to clonal tumors recapitulating the principal clinical, biological, and molecular genetic features of MALT lymphoma. Deletion of p53 gene accelerated tumor onset and induced transformation of MALT lymphoma to activated B-cell diffuse large-cell lymphoma (ABC-DLBCL). Treatment of MALT1-induced lymphomas with a specific inhibitor of MALT1 proteolytic activity decreased cell viability, indicating that endogenous Malt1 signaling was required for tumor cell survival. Our study shows that human-like lymphomas can be modeled in mice by targeting MALT1 expression to hematopoietic stem/progenitor cells, demonstrating the oncogenic role of MALT1 in lymphomagenesis. Furthermore, this work establishes a molecular link between MALT lymphoma and ABC-DLBCL, and provides mouse models to test MALT1 inhibitors. Finally, our results suggest that hematopoietic stem/progenitor cells may be involved in the pathogenesis of human mature B-cell lymphomas. PMID:22689981

  2. Stat3 induces oncogenic Skp2 expression in human cervical carcinoma cells

    SciTech Connect

    Huang, Hanhui; Zhao, Wenrong; Yang, Dan

    2012-02-03

    Highlights: Black-Right-Pointing-Pointer Upregulation of Skp2 by IL-6 or Stat3 activation. Black-Right-Pointing-Pointer Stat3 activates Skp2 expression through bound to its promoter region. Black-Right-Pointing-Pointer Stat3 activates Skp2 expression through recruitment of P300. Black-Right-Pointing-Pointer Stat3 activation decreases the P27 stability. -- Abstract: Dysregulated Skp2 function promotes cell proliferation, which is consistent with observations of Skp2 over-expression in many types of human cancers, including cervical carcinoma (CC). However, the molecular mechanisms underlying elevated Skp2 expression have not been fully explored. Interleukin-6 (IL-6) induced Stat3 activation is viewed as crucial for multiple tumor growth and metastasis. Here, we demonstrate that Skp2 is a direct transcriptional target of Stat3 in the human cervical carcinoma cells. Our data show that IL-6 administration or transfection of a constitutively activated Stat3 in HeLa cells activates Skp2 mRNA transcription. Using luciferase reporter and ChIP assays, we show that Stat3 binds to the promoter region of Skp2 and promotes its activity through recruiting P300. As a result of the increase of Skp2 expression, endogenous p27 protein levels are markedly decreased. Thus, our results suggest a previously unknown Stat3-Skp2 molecular network controlling cervical carcinoma development.

  3. Changes in cortical cytoskeletal and extracellular matrix gene expression in prostate cancer are related to oncogenic ERG deregulation

    PubMed Central

    2010-01-01

    Background The cortical cytoskeleton network connects the actin cytoskeleton to various membrane proteins, influencing cell adhesion, polarity, migration and response to extracellular signals. Previous studies have suggested changes in the expression of specific components in prostate cancer, especially of 4.1 proteins (encoded by EPB41 genes) which form nodes in this network. Methods Expression of EPB41L1, EPB41L2, EPB41L3 (protein: 4.1B), EPB41L4B (EHM2), EPB41L5, EPB49 (dematin), VIL2 (ezrin), and DLG1 (summarized as „cortical cytoskeleton" genes) as well as ERG was measured by quantitative RT-PCR in a well-characterized set of 45 M0 prostate adenocarcinoma and 13 benign tissues. Hypermethylation of EPB41L3 and GSTP1 was compared in 93 cancer tissues by methylation-specific PCR. Expression of 4.1B was further studied by immunohistochemistry. Results EPB41L1 and EPB41L3 were significantly downregulated and EPB41L4B was upregulated in cancer tissues. Low EPB41L1 or high EPB41L4B expression were associated with earlier biochemical recurrence. None of the other cortical cytoskeleton genes displayed expression changes, in particular EPB49 and VIL2, despite hints from previous studies. EPB41L3 downregulation was significantly associated with hypermethylation of its promoter and strongly correlated with GSTP1 hypermethylation. Protein 4.1B was detected most strongly in the basal cells of normal prostate epithelia. Its expression in carcinoma cells was similar to the weaker one in normal luminal cells. EPB41L3 downregulation and EPB41L4B upregulation were essentially restricted to the 22 cases with ERG overexpression. Expression changes in EPB41L3 and EPB41L4B closely paralleled those previously observed for the extracellular matrix genes FBLN1 and SPOCK1, respectively. Conclusions Specific changes in the cortical cytoskeleton were observed during prostate cancer progression. They parallel changes in the expression of extracellular matrix components and all together

  4. Intragenic integration in DLC1 sustains factor VIII expression in primary human cells without insertional oncogenicity

    PubMed Central

    Sivalingam, J; Phan, T T; Kon, O L

    2014-01-01

    Techniques enabling precise genome modifications enhance the safety of gene-based therapy. DLC1 is a hot spot for phiC31 integrase-mediated transgene integration in vitro and in vivo. Here we show that integration of a coagulation factor VIII transgene into intron 7 of DLC1 supports durable expression of factor VIII in primary human umbilical cord-lining epithelial cells. Oligoclonal cells with factor VIII transgene integrated in DLC1 did not have altered expression of DLC1 or neighbouring genes within a 1-Mb interval. Only 1.9% of all expressed genes were transcriptionally altered; most were downregulated and mapped to cell cycle and DNA repair pathways. DLC1-integrated cells were not tumourigenic in vivo and were normal by high-resolution genomic DNA copy number analysis. Our data identify DLC1 as a locus for durable transgene expression that does not incur features of insertional oncogenesis, thus expanding options for developing ex vivo cell therapy mediated by site-specific integration methods. PMID:24553346

  5. [Cell oncogene expression in normal, metaplastic, dysplastic epithelium and squamous cell carcinoma of the uterine cervix].

    PubMed

    Petrov, S V; Mazurenko, N N; Sukhova, N M; Moroz, I P; Katsenel'son, V M; Raĭkhlin, N T; Kiselev, F L

    1994-01-01

    Immunohistochemical analysis of the protein expression c-myc, ets 1, ets 2, TPR-met, c-fos, c-jun, c-ras-pan, p53, yes, src in 79 samples of normal, metaplastic squamous epithelium, intraepithelial and invasive squamous cell carcinoma of uterine cervix was performed using polyclonal rabbit antibodies to the synthetic peptides homologous active areas of corresponding oncoproteins. Higher content of myc, fos, ets2, p53, ras is noted in metaplasia, dysplasia and in tumours as compared to the normal tissues. Protein myc is revealed in the cytoplasm at a grave dysplasia and in the nucleus in the intraepithelial carcinoma: this may serve as a criterion at a differential diagnosis of these conditions. Expression of the oncoproteins fos, ets2, p53, src in the metaplastic squamous cell carcinoma was higher than in the true squamous cell (ectocervical) carcinoma. When compared to the advanced carcinomas, increase of ets2, p53, and at some degree that of myc, the increase is noted in the latter. Invasive carcinoma with a high level of oncoproteins showed a tendency to the synchronization of myc and ras expression. Poor prognosis was associated with a low level (before treatment) of the expression of the majority of the oncoproteins studied. PMID:7848100

  6. Global expression profiling reveals gain-of-function onco-genic activity of a mutated thyroid hormone receptor in thyroid carcinogenesis

    PubMed Central

    Lu, Changxue; Mishra, Alok; Zhu, Yuelin J; Meltzer, Paul; Cheng, Sheue-yann

    2011-01-01

    Thyroid hormone receptors (TRs) are critical in regulating gene expression in normal physiological processes. Decreased expression and/or somatic mutations of TRs have been shown to be associated several types of human cancers including liver, breast, lung, and thyroid. To understand the molecular mechanisms by which mutated TRs promote carcinogenesis, an animal model of follicular thyroid carcinoma (FTC) (Thrbpv/pv mice) was used in the present study. The Thrbpv/pv mouse harbors a knockin dominant negative PV mutation, identified in a patient with resistance to thyroid hormone. To understand whether oncogenic actions of PV involve not only the loss of normal TR functions but also gain-of-function activities, we compared the gene expression profiles of thyroid lesions in Thrbpv/pv mice and Thra1-/- Thrb-/- mice that also spontaneously develop FTC, but with less severe malignancy. Analysis of the cDNA microarray data derived from microdissected thyroid tumor cells of these two mice showed contrasting global gene expression profiles. With stringent selection using 2.5-fold change (p<0.01) in cDNA microarray analysis, 241 genes with altered gene expression were identified. Nearly half of the genes (n=103: 42.7% of total) with altered gene expression in thyroid tumor cells of Thrbpv/pv mice were associated with tumorigenesis and metastasis; some of these genes function as oncogenes in human thyroid cancers. The remaining genes were found to function in transcriptional regulation, RNA processing, cell proliferation, apoptosis, angiogenesis, and cytoskeleton modification. These results indicate that the more aggressive thyroid tumor progression in Thrbpv/pv mice was not due simply to the loss of tumor suppressor functions of TR via mutation but also, importantly, to gain-of-function in the oncogenic activities of PV to drive thyroid carcinogenesis. Thus, the present study identifies a novel mechanism by which a mutated TRβ evolves with an oncogenic advantage to promote

  7. Enforced expression of the c-myc oncogene inhibits cell differentiation by precluding entry into a distinct predifferentiation state in G0/G1.

    PubMed Central

    Freytag, S O

    1988-01-01

    A broad base of data has implicated a role for the c-myc proto-oncogene in the control of the cell cycle and cell differentiation. To further define the role of myc in these processes, I examined the effect of enforced myc expression on several events that are thought to be important steps leading to the terminally differentiated state: (i) the ability to arrest growth in G0/G1, (ii) the ability to replicate the genome upon initiation of the differentiation program, and (iii) the ability to lose responsiveness to mitogens and withdraw from the cell cycle. 3T3-L1 preadipocyte cell lines expressing various levels of myc mRNA were established by transfection with a recombinant myc gene under the transcriptional control of the Rous sarcoma virus (RSV) promoter. Cells that expressed high constitutive levels of pRSVmyc mRNA arrested in G0/G1 at densities similar to those of normal cells at confluence. Upon initiation of the differentiation program, such cells traversed the cell cycle with kinetics similar to those of normal cells and subsequently arrested in G0/G1. Thus, enforced expression of myc had no effect on the ability of cells to arrest growth in G0/G1 or to replicate the genome upon initiation of the differentiation program. Cells were then tested for their ability to reenter the cell cycle upon exposure to high concentrations of serum and for their capacity to differentiate. In contrast to normal cells, cells expressing high constitutive levels of myc RNA reentered the cell cycle when challenged with 30% serum and failed to terminally differentiate. The block to differentiation could be reversed by high expression of myc antisense RNA, showing that the induced block was specifically due to enforced expression of pRSVmyc. These findings indicate that 3T3-L1 preadipocytes enter a specific state in G0/G1 after treatment with differentiation inducers, into which cells expressing high constitutive levels of myc RNA are precluded from entering. I propose that myc

  8. Loss of Dependence on Continued Expression of the Human Papillomavirus 16 E7 Oncogene in Cervical Cancers and Precancerous Lesions Arising in Fanconi Anemia Pathway-Deficient Mice

    PubMed Central

    Park, Soyeong; Park, Jung Wook; Pitot, Henry C.

    2016-01-01

    ABSTRACT   Fanconi anemia (FA) is a rare genetic disorder caused by defects in DNA damage repair. FA patients often develop squamous cell carcinoma (SCC) at sites where high-risk human papillomaviruses (HPVs) are known to cause cancer, including the cervix. However, SCCs found in human FA patients are often HPV negative, even though the majority of female FA patients with anogenital cancers had preexisting HPV-positive dysplasia. We hypothesize that HPVs contribute to the development of SCCs in FA patients but that the continued expression of HPV oncogenes is not required for the maintenance of the cancer state because FA deficiency leads to an accumulation of mutations in cellular genes that render the cancer no longer dependent upon viral oncogenes. We tested this hypothesis, making use of Bi-L E7 transgenic mice in which we temporally controlled expression of HPV16 E7, the dominant viral oncogene in HPV-associated cancers. As seen before, the persistence of cervical neoplastic disease was highly dependent upon the continued expression of HPV16 E7 in FA-sufficient mice. However, in mice with FA deficiency, cervical cancers persisted in a large fraction of the mice after HPV16 E7 expression was turned off, indicating that these cancers had escaped from their dependency on E7. Furthermore, the severity of precancerous lesions also failed to be reduced significantly in the mice with FA deficiency upon turning off expression of E7. These findings confirm our hypothesis and may explain the fact that, while FA patients have a high frequency of infections by HPVs and HPV-induced precancerous lesions, the cancers are frequently HPV negative. Importance   Fanconi anemia (FA) patients are at high risk for developing squamous cell carcinoma (SCC) at sites where high-risk human papillomaviruses (HPVs) frequently cause cancer. Yet these SCCs are often HPV negative. FA patients have a genetic defect in their capacity to repair damaged DNA. HPV oncogenes cause an

  9. Loss of Dependence on Continued Expression of the Human Papillomavirus 16 E7 Oncogene in Cervical Cancers and Precancerous Lesions Arising in Fanconi Anemia Pathway-Deficient Mice.

    PubMed

    Park, Soyeong; Park, Jung Wook; Pitot, Henry C; Lambert, Paul F

    2016-01-01

    Fanconi anemia (FA) is a rare genetic disorder caused by defects in DNA damage repair. FA patients often develop squamous cell carcinoma (SCC) at sites where high-risk human papillomaviruses (HPVs) are known to cause cancer, including the cervix. However, SCCs found in human FA patients are often HPV negative, even though the majority of female FA patients with anogenital cancers had preexisting HPV-positive dysplasia. We hypothesize that HPVs contribute to the development of SCCs in FA patients but that the continued expression of HPV oncogenes is not required for the maintenance of the cancer state because FA deficiency leads to an accumulation of mutations in cellular genes that render the cancer no longer dependent upon viral oncogenes. We tested this hypothesis, making use of Bi-L E7 transgenic mice in which we temporally controlled expression of HPV16 E7, the dominant viral oncogene in HPV-associated cancers. As seen before, the persistence of cervical neoplastic disease was highly dependent upon the continued expression of HPV16 E7 in FA-sufficient mice. However, in mice with FA deficiency, cervical cancers persisted in a large fraction of the mice after HPV16 E7 expression was turned off, indicating that these cancers had escaped from their dependency on E7. Furthermore, the severity of precancerous lesions also failed to be reduced significantly in the mice with FA deficiency upon turning off expression of E7. These findings confirm our hypothesis and may explain the fact that, while FA patients have a high frequency of infections by HPVs and HPV-induced precancerous lesions, the cancers are frequently HPV negative. IMPORTANCE  : Fanconi anemia (FA) patients are at high risk for developing squamous cell carcinoma (SCC) at sites where high-risk human papillomaviruses (HPVs) frequently cause cancer. Yet these SCCs are often HPV negative. FA patients have a genetic defect in their capacity to repair damaged DNA. HPV oncogenes cause an accumulation of DNA

  10. Tipping the balance between good and evil: aberrant 14-3-3ζ expression drives oncogenic TGF-β signaling in metastatic breast cancers.

    PubMed

    Morrison, Chevaun D; Schiemann, William P

    2015-01-01

    Transforming growth factor beta (TGF-β) readily suppresses the development of early-stage breast cancers, an activity that gives way to tumor promotion in their late-stage counterparts. The molecular mechanisms underlying this mysterious switch in TGF-β function remain murky. In addressing this conundrum, Xu et al. observed aberrant 14-3-3ζ expression to prevent the formation of tumor-suppressive Smad2/3:p53 complexes, while simultaneously driving the generation of oncogenic Smad2/3:Gli2 complexes. Once formed, Smad2/3:Gli2 complexes stimulate the expression of parathyroid hormone-related protein necessary for breast cancer metastasis to bone. This viewpoint highlights 14-3-3ζ as an essential driver of oncogenic signaling by Smad2/3 and TGF-β in metastatic breast cancers. PMID:26160166

  11. Perylene and coronene derivatives binding to G-rich promoter oncogene sequences efficiently reduce their expression in cancer cells.

    PubMed

    Micheli, Emanuela; Altieri, Alessandro; Cianni, Lorenzo; Cingolani, Chiara; Iachettini, Sara; Bianco, Armandodoriano; Leonetti, Carlo; Cacchione, Stefano; Biroccio, Annamaria; Franceschin, Marco; Rizzo, Angela

    2016-06-01

    A novel approach to cancer therapeutics is emerging in the field of G-quadruplex (G4) ligands, small molecules designed to stabilize four-stranded structures that can form at telomeres as well as in other genomic sequences, including oncogene promoter sequences, 5'-UTR regions and introns. In this study, we investigated the binding activity of perylene and coronene derivatives PPL3C, CORON and EMICORON to G4 structures formed within the promoter regions of two important cancer-related genes, c-MYC and BCL-2, and their biochemical effects on gene and protein expression. In order to fully characterize the ability of the selected ligands to bind and stabilize the G4 structures originated by the c-MYC and BCL-2 promoter sequences, we performed electrospray ionization mass spectrometry (ESI-MS), Fluorescence Resonance Energy Transfer (FRET) measurements, Circular Dichroism (CD) spectra and polymerase stop assay. Altogether our results showed that the ligands had a high capacity in binding and stabilizing the G4 structures within the c-MYC and BCL-2 promoter sequences in vitro. Notably, when we evaluated by quantitative real-time PCR and western blotting analysis, the effects of treatment with the different G4 ligands on c-MYC and BCL2 expression in a human melanoma cell line, EMICORON appeared the most effective compound in reducing the mRNA and protein levels of both genes. These results encourage to consider EMICORON as a promising example of multimodal class of an antineoplastic drug, affecting different tumor crucial pathways simultaneously: telomere maintenance (as previously described), cell proliferation and apoptosis via down-regulation of both c-MYC and BCL-2 (this paper). PMID:27086081

  12. Low Expression of miR-196b Enhances the Expression of BCR-ABL1 and HOXA9 Oncogenes in Chronic Myeloid Leukemogenesis

    PubMed Central

    Liu, Yue; Zheng, Wenling; Song, Yanbin; Ma, Wenli; Yin, Hong

    2013-01-01

    MicroRNAs (miRNAs) can function as tumor suppressors or oncogene promoters during tumor development. In this study, low levels of expression of miR-196b were detected in patients with chronic myeloid leukemia. Bisulfite genomic sequencing PCR and methylation-specific PCR were used to examine the methylation status of the CpG islands in the miR-196b promoter in K562 cells, patients with leukemia and healthy individuals. The CpG islands showed more methylation in patients with chronic myeloid leukemia compared with healthy individuals (P<0.05), which indicated that low expression of miR-196b may be associated with an increase in the methylation of CpG islands. The dual-luciferase reporter assay system demonstrated that BCR-ABL1 and HOXA9 are the target genes of miR-196b, which was consistent with predictions from bioinformatics software analyses. Further examination of cell function indicated that miR-196b acts to reduce BCR-ABL1 and HOXA9 protein levels, decrease cell proliferation rate and retard the cell cycle. A low level of expression of miR-196b can cause up-regulation of BCR-ABL1 and HOXA9 expression, which leads to the development of chronic myeloid leukemia. MiR-196b may represent an effective target for chronic myeloid leukemia therapy. PMID:23894305

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

    SciTech Connect

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

    1987-03-01

    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.

  14. Cooperative antiproliferative effect of coordinated ectopic expression of DLC1 tumor suppressor protein and silencing of MYC oncogene expression in liver cancer cells: Therapeutic implications

    PubMed Central

    Yang, Xuyu; Zhou, Xiaoling; Tone, Paul; Durkin, Marian E.; Popescu, Nicholas C.

    2016-01-01

    Human hepatocellular carcinoma (HCC) is one of the most common types of cancer and has a very poor prognosis; thus, the development of effective therapies for the treatment of advanced HCC is of high clinical priority. In the present study, the anti-oncogenic effect of combined knockdown of c-Myc expression and ectopic restoration of deleted in liver cancer 1 (DLC1) expression was investigated in human liver cancer cells. Expression of c-Myc in human HCC cells was knocked down by stable transfection with a Myc-specific short hairpin (sh) RNA vector. DLC1 expression in Huh7 cells was restored by adenovirus transduction, and the effects of DLC1 expression and c-Myc knockdown on Ras homolog gene family, member A (RhoA) levels, cell proliferation, soft agar colony formation and cell invasion were measured. Downregulation of c-Myc or re-expression of DLC1 led to a marked reduction in RhoA levels, which was associated with decreases in cell proliferation, soft agar colony formation and invasiveness; this inhibitory effect was augmented with a combination of DLC1 transduction and c-Myc suppression. To determine whether liver cell-specific delivery of DLC1 was able to enhance the inhibitory effect of c-Myc knockdown on tumor growth in vivo, DLC1 vector DNA complexed with galactosylated polyethylene glycol-linear polyethyleneimine was administered by tail vein injection to mice bearing subcutaneous xenografts of Huh7 cells transfected with shMyc or control shRNA. A cooperative inhibitory effect of DLC1 expression and c-Myc knockdown on the growth of Huh7-derived tumors was observed, suggesting that targeted liver cell delivery of DLC1 and c-Myc shRNA may serve as a possible gene therapy modality for the treatment of human HCC. PMID:27446476

  15. Preventive efficacy of receptor class selective retinoids on HER-2/neu oncogene expressing preneoplastic human mammary epithelial cells.

    PubMed

    Jinno, Hiromitsu; Steiner, Melissa G; Nason-Burchenal, Kathryn; Osborne, Michael P; Telang, Nitin T

    2002-07-01

    Aberrant proliferation is an early-occurring event in vitro prior to tumorigenesis in vivo in the multistep process of carcinogenesis. Inhibition of aberrant proliferation therefore may represent a useful biomarker to evaluate the efficacy of chemopreventive agents. Retinoids have exhibited preventive efficacy in vitro and in vivo predominantly through the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs). Clinically relevant biochemical and cellular mechanistic endpoints for chemopreventive effects of retinoids should provide novel biomarkers. The present study was designed to examine the preventive efficacy of natural retinoids, all-trans-retinoic acid (ATRA) and 9-cis-retinoic acid (9cisRA), and to identify the possible mechanisms for their effects using the HER-2/neu oncogene expressing preneoplastic human mammary epithelial 184-B5/HER cells. Seven-day treatment with ATRA and 9cisRA exhibited a dose-dependent growth inhibition. Long-term (21 days) treatment with IC20 doses of 50 nM ATRA and 100 nM 9cisRA inhibited anchorage-dependent colony forming efficiency by about 75.4% (p<0.01) and 84.9% (p<0.01), respectively. Cell cycle analysis revealed that a 24-h treatment with IC90 doses of 2 microM ATRA and 3 microM 9cisRA accumulates cells in the G0/G1 phase and inhibit S and/or G2/M phase of the cell cycle. ATRA and 9cisRA induced an 11-fold (p=0.03) and a 9-fold (p=0.04) increase in subG0/G1 (apoptotic) population relative to the solvent control, respectively. ATRA and 9cisRA induced 77% (p=0.01) and 51% (p=0.02) decrease in tyrosine kinase immunoreactivity, respectively. Similarly, the two retinoids caused almost a 50% (p=0.01) down-regulation of Bcl-2 immunoreactivity. Western blot analysis revealed that ATRA induced an increase in RARbeta expression and a decrease in RARgamma expression, while 9cisRA down-regulated RXRalpha expression. These data demonstrate that ATRA and 9cisRA may inhibit HER-2/neu induced aberrant proliferation in part by

  16. Induction of cell death by stimulation of protein kinase C in human epithelial cells expressing a mutant ras oncogene: a potential therapeutic target.

    PubMed Central

    Hall-Jackson, C. A.; Jones, T.; Eccles, N. G.; Dawson, T. P.; Bond, J. A.; Gescher, A.; Wynford-Thomas, D.

    1998-01-01

    Ras oncogene activation is a key genetic event in several types of human cancer, making its signal pathways an ideal target for novel therapies. We previously showed that expression of mutant ras sensitizes human thyroid epithelial cells to induction of cell death by treatment with phorbol 12-myristate 13-acetate (PMA) and other phorbol esters. We have now investigated further the nature and mechanism of this cell death using both primary and cell line models. The cytotoxic effect of PMA could be blocked by bisindolylmaleimide (GF 109203X), a well-characterized inhibitor of c and n protein kinase C (PKC) isoforms, and by prior down-regulation of PKC, indicating that it is mediated by acute stimulation, rather than down-regulation. Western analysis identified two candidate isoforms--alpha and epsilon--both of which showed PMA-induced subcellular translocation, either or both of which may be necessary for PMA-induced cell death. Immunofluorescence showed that PMA induced a rapid nuclear translocation of p42 MAP kinase of similar magnitude in the presence or absence of mutant ras expression. Cell death exhibited the microscopic features (chromatin condensation, TdT labelling) and DNA fragmentation typical of apoptosis but after a surprising lag (4 days). Taken together with recent models of ras-modulated apoptosis, our data suggest that activation of the MAPK pathway by PMA tips the balance of pro- and anti-apoptotic signals generated by ras in favour of apoptosis. The high frequency of ras mutations in some cancers, such as cancer of the pancreas, which are refractory to conventional chemotherapy, together with the potential for stimulating PKC by cell-permeant pharmacological agents, makes this an attractive therapeutic approach. Images Figure 1 Figure 2 Figure 4 Figure 5 Figure 7 Figure 6 Figure 8 PMID:9744505

  17. Repression of CD24 surface protein expression by oncogenic Ras is relieved by inhibition of Raf but not MEK or PI3K

    PubMed Central

    Pallegar, Nikitha K.; Ayre, D. Craig; Christian, Sherri L.

    2015-01-01

    CD24 is a dynamically regulated cell surface protein. High expression of CD24 leads to progression of lung, prostrate, colon, and pancreatic cancers, among others. In contrast, low expression of CD24 leads to cell proliferation and metastasis of breast cancer stem cells (BCSCs). Activating mutations in Ras are found in 30% of all human cancers. Oncogenic Ras constitutively stimulates the Raf, PI3K, and Ral GDS signaling pathways, leading to cellular transformation. Previous studies have shown that expression of oncogenic Ras in breast cancer cells generates CD24− cells from CD24+ cells. However, the molecular mechanisms involved in the generation of CD24− cells were not determined. Here, we demonstrate that oncogenic Ras (RasV12) expression suppresses CD24 mRNA, protein, and promoter levels when expressed in NIH/3T3 cells. Furthermore, activation of only the Raf pathway was sufficient to downregulate CD24 mRNA and protein expression to levels similar to those seen in with RasV12 expression. In contrast, activation of the PI3K pathway downregulated mRNA expression with a partial effect on protein expression whereas activation of the RalGDS pathway only partially affected protein expression. Surprisingly, inhibition of MEK with U0126 only partially restored CD24 mRNA expression but not surface protein expression. In contrast, inhibition of Raf with sorafenib did not restore CD24 mRNA expression but significantly increased the proportion of RasV12 cells expressing CD24. Therefore, the Raf pathway is the major repressor of CD24 mRNA and protein expression, with PI3K also able to substantially inhibit CD24 expression. Moreover, these data indicate that the levels of CD24 mRNA and surface protein are independently regulated. Although inhibition of Raf by sorafenib only partially restored CD24 expression, sorafenib should still be considered as a potential therapeutic strategy to alter CD24 expression in CD24− cells, such as BCSCs. PMID:26301220

  18. Suppression of protein kinase C and nuclear oncogene expression as possible action mechanisms of cancer chemoprevention by Curcumin.

    PubMed

    Lin, Jen-Kun

    2004-07-01

    Curcumin (diferuloylmethane) is a major naturally-occurring polyphenol of Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. Curcumin has shown anti-carcinogenic activity in animal models. Curcumin possesses anti-inflammatory activity and is a potent inhibitor of reactive oxygen-generating enzymes such as lipoxygenase/cyclooxygenase, xanthine dehydrogenase/oxidase and inducible nitric oxide synthase; and an effective inducer of heme oxygenase-1. Curcumin is also a potent inhibitor of protein kinase C (PKC), EGF(Epidermal growth factor)-receptor tyrosine kinase and IkappaB kinase. Subsequently, curcumin inhibits the activation of NF(nucleor factor)kappaB and the expressions of oncogenes including c-jun, c-fos, c-myc, NIK, MAPKs, ERK, ELK, PI3K, Akt, CDKs and iNOS. It is proposed that curcumin may suppress tumor promotion through blocking signal transduction pathways in the target cells. The oxidant tumor promoter TPA activates PKC by reacting with zinc thiolates present within the regulatory domain, while the oxidized form of cancer chemopreventive agent such as curcumin can inactivate PKC by oxidizing the vicinal thiols present within the catalytic domain. Recent studies indicated that proteasome-mediated degradation of cell proteins play a pivotal role in the regulation of several basic cellular processes including differentiation, proliferation, cell cycling, and apoptosis. It has been demonstrated that curcumin-induced apoptosis is mediated through the impairment of ubiquitin-proteasome pathway. Curcumin was first biotransformed to dihydrocurcumin and tetrahydrocurcumin and that these compounds subsequently were converted to monoglucuronide conjugates. These results suggest that curcumin-glucuronide, dihydrocurcumin-glucuronide, tetrahydrocurcumin-glucuronide and tetrahydrocurcumin are the major metabolites of curcumin in mice, rats and humans. PMID:15356994

  19. Development of a conditional liver tumor model by mifepristone-inducible Cre recombination to control oncogenic krasV12 expression in transgenic zebrafish

    PubMed Central

    Nguyen, Anh Tuan; Koh, Vivien; Spitsbergen, Jan M.; Gong, Zhiyuan

    2016-01-01

    Here we report a new transgenic expression system by combination of liver-specific expression, mifepristone induction and Cre-loxP recombination to conditionally control the expression of oncogenic krasV12. This transgenic system allowed expression of krasV12 specifically in the liver by a brief exposure of mifepristone to induce permanent genomic recombination mediated by the Cre-loxP system. We found that liver tumors were generally induced from multiple foci due to incomplete Cre-loxP recombination, thus mimicking naturally occurring human tumors resulting from one or a few mutated cells and clonal proliferation to form nodules. Similar to our earlier studies by both constitutive and inducible expression of the krasV12 oncogene, hepatocellular carcinoma (HCC) is the main type of liver tumor induced by krasV12 expression. Moreover, mixed tumors with hepatocellular adenoma and hepatoblastoma (HB) were also frequently observed. Molecular analyses also indicated similar increase of phosphorylated ERK1/2 in all types of liver tumors, but nuclear localization of β–catenin, a sign of malignant transformation, was found only in HCC and HB. Taken together, our new transgenic system reported in this study allows transgenic krasV12 expression specifically in the zebrafish liver only by a brief exposure of mifepristone to induce permanent genomic recombination mediated by the Cre-loxP system. PMID:26790949

  20. Prevention of tumor growth driven by PIK3CA and HPV oncogenes by targeting mTOR signaling with metformin in oral squamous carcinomas expressing OCT3

    PubMed Central

    Madera, Dmitri; Vitale-Cross, Lynn; Martin, Daniel; Schneider, Abraham; Molinolo, Alfredo A.; Gangane, Nitin; Carey, Thomas E.; McHugh, Jonathan B.; Komarck, Christine M.; Walline, Heather M.; William, William N.; Seethala, Raja R.; Ferris, Robert; Gutkind, J. Silvio

    2015-01-01

    Most head and neck squamous cell carcinomas (HNSCC) exhibit a persistent activation of the PI3K-mTOR signaling pathway. We have recently shown that metformin, an oral antidiabetic drug that is also used to treat lipodystrophy in HIV-infected (HIV+) individuals, diminishes mTOR activity and prevents the progression of chemically-induced experimental HNSCC premalignant lesions. Here, we explored the preclinical activity of metformin in HNSCCs harboring PIK3CA mutations and HPV oncogenes, both representing frequent HNSCC alterations, aimed at developing effective targeted preventive strategies. The biochemical and biological effects of metformin were evaluated in representative HNSCC cells expressing mutated PIK3CA or HPV oncogenes (HPV+). The oral delivery of metformin was optimized to achieve clinical relevant blood levels. Molecular determinants of metformin sensitivity were also investigated, and their expression levels examined in a large collection of HNSCC cases. We found that metformin inhibits mTOR signaling and tumor growth in HNSCC cells expressing mutated PIK3CA and HPV oncogenes, and that these activities require the expression of organic cation transporter 3 (OCT3/SLC22A3), a metformin uptake transporter. Co-expression of OCT3 and the mTOR pathway activation marker pS6 were observed in most HNSCC cases, including those arising in HIV+ patients. Activation of the PI3K-mTOR pathway is a widespread event in HNSCC, including HPV− and HPV+ lesions arising in HIV+ patients, all of which co-express OCT3. These observations may provide a rationale for the clinical evaluation of metformin to halt HNSCC development from precancerous lesions, including in HIV+ individuals at risk of developing HPV-associated cancers. PMID:25681087

  1. The Jak2V617F oncogene associated with myeloproliferative diseases requires a functional FERM domain for transformation and for expression of the Myc and Pim proto-oncogenes

    PubMed Central

    Wernig, Gerlinde; Gonneville, Jeffrey R.; Crowley, Brian J.; Rodrigues, Margret S.; Reddy, Mamatha M.; Hudon, Heidi E.; Walz, Christoph; Reiter, Andreas; Podar, Klaus; Royer, Yohan; Constantinescu, Stefan N.; Tomasson, Michael H.; Griffin, James D.; Gilliland, D. Gary

    2008-01-01

    The V617F activating point mutation in Jak2 is associated with a proportion of myeloproliferative disorders. In normal hematopoietic cells, Jak2 signals only when associated with a growth factor receptor, such as the erythropoietin receptor (EpoR). We sought to identify the molecular requirements for activation of Jak2V617F by introducing a point mutation in the FERM domain (Y114A), required for receptor binding. Whereas BaF3.EpoR cells are readily transformed by Jak2V617F to Epo independence, we found that the addition of the FERM domain mutation blocked transformation and the induction of reactive oxygen species. Further, while cells expressing Jak2V617F had constitutive activation of STAT5, cells expressing Jak2V617F/Y114A did not, suggesting that signaling is defective at a very proximal level. In addition, expression of the Myc and Pim proto-oncogenes by Jak2V617F was found to be FERM domain dependent. An inducible constitutively active STAT5 mutant expressed in BaF3 cells was sufficient to induce Myc and Pim. Finally, the FERM domain in Jak2V617F was also required for abnormal hematopoiesis in transduced primary murine fetal liver cells. Overall, our results suggest that constitutive activation of Jak2 requires an intact FERM domain for a transforming phenotype, and is necessary for activation of the major target of Jak2, STAT5. PMID:18216297

  2. Prevention of tumor growth driven by PIK3CA and HPV oncogenes by targeting mTOR signaling with metformin in oral squamous carcinomas expressing OCT3.

    PubMed

    Madera, Dmitri; Vitale-Cross, Lynn; Martin, Daniel; Schneider, Abraham; Molinolo, Alfredo A; Gangane, Nitin; Carey, Thomas E; McHugh, Jonathan B; Komarck, Christine M; Walline, Heather M; William, William N; Seethala, Raja R; Ferris, Robert L; Gutkind, J Silvio

    2015-03-01

    Most squamous cell carcinomas of the head and neck (HNSCC) exhibit a persistent activation of the PI3K-mTOR signaling pathway. We have recently shown that metformin, an oral antidiabetic drug that is also used to treat lipodystrophy in HIV-infected (HIV(+)) individuals, diminishes mTOR activity and prevents the progression of chemically induced experimental HNSCC premalignant lesions. Here, we explored the preclinical activity of metformin in HNSCCs harboring PIK3CA mutations and HPV oncogenes, both representing frequent HNSCC alterations, aimed at developing effective targeted preventive strategies. The biochemical and biologic effects of metformin were evaluated in representative HNSCC cells expressing mutated PIK3CA or HPV oncogenes (HPV(+)). The oral delivery of metformin was optimized to achieve clinical relevant blood levels. Molecular determinants of metformin sensitivity were also investigated, and their expression levels were examined in a large collection of HNSCC cases. We found that metformin inhibits mTOR signaling and tumor growth in HNSCC cells expressing mutated PIK3CA and HPV oncogenes, and that these activities require the expression of organic cation transporter 3 (OCT3/SLC22A3), a metformin uptake transporter. Coexpression of OCT3 and the mTOR pathway activation marker pS6 were observed in most HNSCC cases, including those arising in HIV(+) patients. Activation of the PI3K-mTOR pathway is a widespread event in HNSCC, including HPV(-) and HPV(+) lesions arising in HIV(+) patients, all of which coexpress OCT3. These observations may provide a rationale for the clinical evaluation of metformin to halt HNSCC development from precancerous lesions, including in HIV(+) individuals at risk of developing HPV(-) associated cancers. PMID:25681087

  3. Soy isoflavone genistein modulates cell cycle progression and induces apoptosis in HER-2/neu oncogene expressing human breast epithelial cells.

    PubMed

    Katdare, Meena; Osborne, Michael; Telang, Nitin T

    2002-10-01

    In the multistep progressive pathogenesis of human breast cancer, comedo ductal carcinoma in situ (DCIS) represents a preinvasive precursor lesion for therapy resistant invasive cancer. Human tissue derived cell culture models exhibiting molecular similarities to clinical DCIS facilitate an important preclinical mechanistic approach for evaluation of preventive efficacy of natural and synthetic chemopreventive compounds. Natural phytochemicals present in fresh fruits, vegetables and grain products are likely to offer protection against cancer. The clinical efficacy of these natural phytochemicals, however, depends on extrapolation, and is therefore equivocal. The present study determined whether the natural soy isoflavone genistein (GEN) inhibited aberrant proliferation in 184-B5/HER cells (a model for human comedo DCIS) and identified possible mechanisms responsible for its efficacy. Human reduction mammoplasty derived HER-2/neu oncogene expressing preneoplastic 184-B5/HER cells represented the experimental system. Flow cytometry and cellular epifluorescence based assays were utilized to quantitate the alterations in cell cycle progression, cellular apoptosis, and in the status of cell cycle regulatory and apoptosis-associated gene product expression. The 184-B5/HER cells exhibited specific immunofluorescence to p185HER, p53, EGFR, but not to ERalpha, thus resembling comedo DCIS. Treatment of 184-B5/HER cells with GEN resulted in a dose-dependent decrease in the viable cell population, increase in the G0/G1:S + G2/M ratio and enhancement of sub G0/G1 (apoptotic population). Exposure to the maximum cytostatic 10 microM dose of GEN down-regulated HER-2/neu mediated signal transduction as evidenced by a 73.9% decrease (p=0.001) in p185HER specific, and a 89.8% decrease (p=0.001) in phosphotyrosine specific immunofluorescence. The increase in G0/G1:S + G2/M ratio in response to the treatment with 10 microM GEN was associated with a 85.5% decrease (p=0.001) in

  4. Global gene expression changes of in vitro stimulated human transformed germinal centre B cells as surrogate for oncogenic pathway activation in individual aggressive B cell lymphomas

    PubMed Central

    2012-01-01

    Background Aggressive Non-Hodgkin lymphomas (NHL) are a group of lymphomas derived from germinal centre B cells which display a heterogeneous pattern of oncogenic pathway activation. We postulate that specific immune response associated signalling, affecting gene transcription networks, may be associated with the activation of different oncogenic pathways in aggressive Non-Hodgkin lymphomas (NHL). Methodology The B cell receptor (BCR), CD40, B-cell activating factor (BAFF)-receptors and Interleukin (IL) 21 receptor and Toll like receptor 4 (TLR4) were stimulated in human transformed germinal centre B cells by treatment with anti IgM F(ab)2-fragments, CD40L, BAFF, IL21 and LPS respectively. The changes in gene expression following the activation of Jak/STAT, NF-кB, MAPK, Ca2+ and PI3K signalling triggered by these stimuli was assessed using microarray analysis. The expression of top 100 genes which had a change in gene expression following stimulation was investigated in gene expression profiles of patients with Aggressive non-Hodgkin Lymphoma (NHL). Results αIgM stimulation led to the largest number of changes in gene expression, affecting overall 6596 genes. While CD40L stimulation changed the expression of 1194 genes and IL21 stimulation affected 902 genes, only 283 and 129 genes were modulated by lipopolysaccharide or BAFF receptor stimulation, respectively. Interestingly, genes associated with a Burkitt-like phenotype, such as MYC, BCL6 or LEF1, were affected by αIgM. Unique and shared gene expression was delineated. NHL-patients were sorted according to their similarity in the expression of TOP100 affected genes to stimulated transformed germinal centre B cells The αIgM gene module discriminated individual DLBCL in a similar manner to CD40L or IL21 gene modules. DLBCLs with low module activation often carry chromosomal MYC aberrations. DLBCLs with high module activation show strong expression of genes involved in cell-cell communication, immune responses

  5. neu protooncogene fused to an immunoglobulin heavy chain gene requires immunoglobulin light chain for cell surface expression and oncogenic transformation.

    PubMed Central

    Flanagan, J G; Leder, P

    1988-01-01

    The protein encoded by the neu protooncogene (human gene symbol NGL for neuro/glioblastoma-derived) is a member of the surface receptor/tyrosine kinase family. Though its structure suggests that it can transduce a transmembrane signal, neither its extracellular ligand nor its critical intracellular substrates are known. To explore the functional properties of the protein encoded by neu, we created a fusion gene that joins the cytoplasmic domain of neu to the extracellular portion of an immunoglobulin heavy chain. The localization of the fusion polypeptide can then be controlled by coexpression with immunoglobulin light chain. In the absence of light chain, the heavy chain-neu polypeptide is expressed intracellularly and has no transforming activity. By contrast, in the presence of light chain the fusion polypeptide is expressed at the cell surface and produces tumorigenic foci. Thus, transformation apparently requires expression at the cell surface, where the neu intracellular domain can interact with components that are localized to the plasma membrane. The fusion protein is active in cellular transformation when the transmembrane domain is derived either from neu or from immunoglobulin, indicating that the neu transmembrane domain is not specifically required for transformation, although neu activation in tumors is known to result from a point mutation in this region. The extracellular immunoglobulin heavy and light chain domains of the fusion protein form a functional binding site that allows antigen to modulate its activity, reversing the transforming effect. Images PMID:2903500

  6. Acidosis decreases c-Myc oncogene expression in human lymphoma cells: a role for the proton-sensing G protein-coupled receptor TDAG8.

    PubMed

    Li, Zhigang; Dong, Lixue; Dean, Eric; Yang, Li V

    2013-01-01

    Acidosis is a biochemical hallmark of the tumor microenvironment. Here, we report that acute acidosis decreases c-Myc oncogene expression in U937 human lymphoma cells. The level of c-Myc transcripts, but not mRNA or protein stability, contributes to c-Myc protein reduction under acidosis. The pH-sensing receptor TDAG8 (GPR65) is involved in acidosis-induced c-Myc downregulation. TDAG8 is expressed in U937 lymphoma cells, and the overexpression or knockdown of TDAG8 further decreases or partially rescues c-Myc expression, respectively. Acidic pH alone is insufficient to reduce c-Myc expression, as it does not decrease c-Myc in H1299 lung cancer cells expressing very low levels of pH-sensing G protein-coupled receptors (GPCRs). Instead, c-Myc is slightly increased by acidosis in H1299 cells, but this increase is completely inhibited by ectopic overexpression of TDAG8. Interestingly, TDAG8 expression is decreased by more than 50% in human lymphoma samples in comparison to non-tumorous lymph nodes and spleens, suggesting a potential tumor suppressor function of TDAG8 in lymphoma. Collectively, our results identify a novel mechanism of c-Myc regulation by acidosis in the tumor microenvironment and indicate that modulation of TDAG8 and related pH-sensing receptor pathways may be exploited as a new approach to inhibit Myc expression. PMID:24152439

  7. Acidosis Decreases c-Myc Oncogene Expression in Human Lymphoma Cells: A Role for the Proton-Sensing G Protein-Coupled Receptor TDAG8

    PubMed Central

    Li, Zhigang; Dong, Lixue; Dean, Eric; Yang, Li V.

    2013-01-01

    Acidosis is a biochemical hallmark of the tumor microenvironment. Here, we report that acute acidosis decreases c-Myc oncogene expression in U937 human lymphoma cells. The level of c-Myc transcripts, but not mRNA or protein stability, contributes to c-Myc protein reduction under acidosis. The pH-sensing receptor TDAG8 (GPR65) is involved in acidosis-induced c-Myc downregulation. TDAG8 is expressed in U937 lymphoma cells, and the overexpression or knockdown of TDAG8 further decreases or partially rescues c-Myc expression, respectively. Acidic pH alone is insufficient to reduce c-Myc expression, as it does not decrease c-Myc in H1299 lung cancer cells expressing very low levels of pH-sensing G protein-coupled receptors (GPCRs). Instead, c-Myc is slightly increased by acidosis in H1299 cells, but this increase is completely inhibited by ectopic overexpression of TDAG8. Interestingly, TDAG8 expression is decreased by more than 50% in human lymphoma samples in comparison to non-tumorous lymph nodes and spleens, suggesting a potential tumor suppressor function of TDAG8 in lymphoma. Collectively, our results identify a novel mechanism of c-Myc regulation by acidosis in the tumor microenvironment and indicate that modulation of TDAG8 and related pH-sensing receptor pathways may be exploited as a new approach to inhibit Myc expression. PMID:24152439

  8. Oncogene activation and tumor suppressor gene inactivation find their sites of expression in the changes in time and space of the age-adjusted cancer incidence rate.

    PubMed

    Kodama, M; Kodama, T; Murakami, M

    2000-01-01

    profile in which the correlation coefficient r, a measure of fitness to the 2 equilibrium models, is converted to either +(r > 0) or -(0 > r) for each of the original-, the Rect-, and the Para-coordinates was found to be informative in identifying a group of tumors with sex discrimination of cancer risk (log AAIR changes in space) or another group of environmental hormone-linked tumors (log AAIR changes in time and space)--a finding to indicate that the r-profile of a given tumor, when compared with other neoplasias, may provide a clue to investigating the biological behavior of the tumor. 4) The recent risk increase of skin cancer of both sexes, being classified as an example of environmental hormone-linked neoplasias, was found to commit its ascension of cancer risk along the direction of the centrifugal forces of the time- and space-linked tumor suppressor gene inactivation plotted in the 2-dimension diagram. In conclusion, the centripetal force of oncogene activation and centrifugal force of tumor suppressor gene inactivation found their sites of expression in the distribution pattern of a cancer risk parameter, log AAIR, of a given neoplasias of both sexes on the 2-dimension diagram. The application of the least square method of Gauss to the log AAIR changes in time and space, and also with and without topological modulations of the original sets, when presented in terms of the r-profile, was found to be informative in understanding behavioral characteristics of human neoplaisias. PMID:11204489

  9. Tobacco exposure results in increased E6 and E7 oncogene expression, DNA damage and mutation rates in cells maintaining episomal human papillomavirus 16 genomes

    PubMed Central

    Wei, Lanlan; Griego, Anastacia M.; Chu, Ming; Ozbun, Michelle A.

    2014-01-01

    High-risk human papillomavirus (HR-HPV) infections are necessary but insufficient agents of cervical and other epithelial cancers. Epidemiological studies support a causal, but ill-defined, relationship between tobacco smoking and cervical malignancies. In this study, we used mainstream tobacco smoke condensate (MSTS-C) treatments of cervical cell lines that maintain either episomal or integrated HPV16 or HPV31 genomes to model tobacco smoke exposure to the cervical epithelium of the smoker. MSTS-C exposure caused a dose-dependent increase in viral genome replication and correspondingly higher early gene transcription in cells with episomal HPV genomes. However, MSTS-C exposure in cells with integrated HR-HPV genomes had no effect on genome copy number or early gene transcription. In cells with episomal HPV genomes, the MSTS-C-induced increases in E6 oncogene transcription led to decreased p53 protein levels and activity. As expected from loss of p53 activity in tobacco-exposed cells, DNA strand breaks were significantly higher but apoptosis was minimal compared with cells containing integrated viral genomes. Furthermore, DNA mutation frequencies were higher in surviving cells with HPV episomes. These findings provide increased understanding of tobacco smoke exposure risk in HPV infection and indicate tobacco smoking acts more directly to alter HR-HPV oncogene expression in cells that maintain episomal viral genomes. This suggests a more prominent role for tobacco smoke in earlier stages of HPV-related cancer progression. PMID:25064354

  10. v-cbl, an oncogene from a dual-recombinant murine retrovirus that induces early B-lineage lymphomas.

    PubMed Central

    Langdon, W Y; Hartley, J W; Klinken, S P; Ruscetti, S K; Morse, H C

    1989-01-01

    Cas NS-1 is an acutely transforming murine retrovirus that induces pre-B and pro-B cell lymphomas. Molecular cloning showed it was generated from the ecotropic Cas-Br-M virus by sequential recombinations with endogenous retroviral sequences and a cellular oncogene. The oncogene sequence shows no homology with known oncogenes but some similarity to the yeast transcriptional activator GCN4. A 100-kDa gag-cbl fusion protein, with no detectable kinase activity, is responsible for the cellular transformation. The cellular homologue of v-cbl, present in mouse and human DNA, is expressed in a range of hemopoietic lineages. Images PMID:2784003

  11. Modulation of EZH2 Expression by MEK-ERK or PI3K-AKT Signaling in Lung Cancer Is Dictated by Different KRAS Oncogene Mutations.

    PubMed

    Riquelme, Erick; Behrens, Carmen; Lin, Heather Y; Simon, George; Papadimitrakopoulou, Vassiliki; Izzo, Julie; Moran, Cesar; Kalhor, Neda; Lee, J Jack; Minna, John D; Wistuba, Ignacio I

    2016-02-01

    EZH2 overexpression promotes cancer by increasing histone methylation to silence tumor suppressor genes, but how EZH2 levels become elevated in cancer is not understood. In this study, we investigated the mechanisms by which EZH2 expression is regulated in non-small cell lung carcinoma cells by oncogenic KRAS. In cells harboring KRAS(G12C) and KRAS(G12D) mutations, EZH2 expression was modulated by MEK-ERK and PI3K/AKT signaling, respectively. Accordingly, MEK-ERK depletion decreased EZH2 expression in cells harboring the KRAS(G12C) mutation, whereas PI3K/AKT depletion decreased EZH2 expression, EZH2 phosphorylation, and STAT3 activity in KRAS(G12D)-mutant cell lines. Combined inhibition of EZH2 and MEK-ERK or PI3K/AKT increased the sensitivity of cells with specific KRAS mutations to MEK-ERK and PI3K/AKT-targeted therapies. Our work defines EZH2 as a downstream effector of KRAS signaling and offers a rationale for combining EZH2 inhibitory strategies with MEK-ERK- or PI3K/AKT-targeted therapies to treat lung cancer patients, as stratified into distinct treatment groups based on specific KRAS mutations. PMID:26676756

  12. Expression of an Oncogenic BARD1 Splice Variant Impairs Homologous Recombination and Predicts Response to PARP-1 Inhibitor Therapy in Colon Cancer

    PubMed Central

    Ozden, Ozkan; Bishehsari, Faraz; Bauer, Jessica; Park, Seong-Hoon; Jana, Arundhati; Baik, Seung Hyun; Sporn, Judith C.; Staudacher, Jonas J.; Yazici, Cemal; Krett, Nancy; Jung, Barbara

    2016-01-01

    BRCA1-associated RING domain protein 1 (BARD1) stabilizes BRCA1 protein by forming a heterodimeric RING-RING complex, and impacts function of BRCA1, including homologous recombination (HR) repair. Although colon cancer cells usually express wild type BRCA1, presence of an oncogenic BARD1 splice variant (SV) in select cancers may render BRCA1 dysfunctional and allow cells to become sensitive to HR targeting therapies. We previously reported association of loss of full-length (FL) BARD1 with poor prognosis in colon cancer as well as expression of various BARD1 SVs with unknown function. Here we show that loss of BARD1 function through the expression of a BARD1 SV, BARD1β, results in a more malignant phenotype with decreased RAD51 foci formation, reduced BRCA1 E3 ubiquitin ligase activity, and decreased nuclear BRCA1 protein localization. BARD1β sensitizes colon cancer cells to poly ADP ribose polymerase 1 (PARP-1) inhibition even in a FL BRCA1 background. These results suggest that expression of BARD1β may serve as a future biomarker to assess suitability of colon cancers for HR targeting with PARP-1 inhibitors in treatment of advanced colon cancer. PMID:27197561

  13. Expression of an Oncogenic BARD1 Splice Variant Impairs Homologous Recombination and Predicts Response to PARP-1 Inhibitor Therapy in Colon Cancer.

    PubMed

    Ozden, Ozkan; Bishehsari, Faraz; Bauer, Jessica; Park, Seong-Hoon; Jana, Arundhati; Baik, Seung Hyun; Sporn, Judith C; Staudacher, Jonas J; Yazici, Cemal; Krett, Nancy; Jung, Barbara

    2016-01-01

    BRCA1-associated RING domain protein 1 (BARD1) stabilizes BRCA1 protein by forming a heterodimeric RING-RING complex, and impacts function of BRCA1, including homologous recombination (HR) repair. Although colon cancer cells usually express wild type BRCA1, presence of an oncogenic BARD1 splice variant (SV) in select cancers may render BRCA1 dysfunctional and allow cells to become sensitive to HR targeting therapies. We previously reported association of loss of full-length (FL) BARD1 with poor prognosis in colon cancer as well as expression of various BARD1 SVs with unknown function. Here we show that loss of BARD1 function through the expression of a BARD1 SV, BARD1β, results in a more malignant phenotype with decreased RAD51 foci formation, reduced BRCA1 E3 ubiquitin ligase activity, and decreased nuclear BRCA1 protein localization. BARD1β sensitizes colon cancer cells to poly ADP ribose polymerase 1 (PARP-1) inhibition even in a FL BRCA1 background. These results suggest that expression of BARD1β may serve as a future biomarker to assess suitability of colon cancers for HR targeting with PARP-1 inhibitors in treatment of advanced colon cancer. PMID:27197561

  14. Dependence of Intracellular and Exosomal microRNAs on Viral E6/E7 Oncogene Expression in HPV-positive Tumor Cells

    PubMed Central

    Honegger, Anja; Schilling, Daniela; Bastian, Sandra; Sponagel, Jasmin; Kuryshev, Vladimir; Sültmann, Holger; Scheffner, Martin; Hoppe-Seyler, Karin; Hoppe-Seyler, Felix

    2015-01-01

    Specific types of human papillomaviruses (HPVs) cause cervical cancer. Cervical cancers exhibit aberrant cellular microRNA (miRNA) expression patterns. By genome-wide analyses, we investigate whether the intracellular and exosomal miRNA compositions of HPV-positive cancer cells are dependent on endogenous E6/E7 oncogene expression. Deep sequencing studies combined with qRT-PCR analyses show that E6/E7 silencing significantly affects ten of the 52 most abundant intracellular miRNAs in HPV18-positive HeLa cells, downregulating miR-17-5p, miR-186-5p, miR-378a-3p, miR-378f, miR-629-5p and miR-7-5p, and upregulating miR-143-3p, miR-23a-3p, miR-23b-3p and miR-27b-3p. The effects of E6/E7 silencing on miRNA levels are mainly not dependent on p53 and similarly observed in HPV16-positive SiHa cells. The E6/E7-regulated miRNAs are enriched for species involved in the control of cell proliferation, senescence and apoptosis, suggesting that they contribute to the growth of HPV-positive cancer cells. Consistently, we show that sustained E6/E7 expression is required to maintain the intracellular levels of members of the miR-17~92 cluster, which reduce expression of the anti-proliferative p21 gene in HPV-positive cancer cells. In exosomes secreted by HeLa cells, a distinct seven-miRNA-signature was identified among the most abundant miRNAs, with significant downregulation of let-7d-5p, miR-20a-5p, miR-378a-3p, miR-423-3p, miR-7-5p, miR-92a-3p and upregulation of miR-21-5p, upon E6/E7 silencing. Several of the E6/E7-dependent exosomal miRNAs have also been linked to the control of cell proliferation and apoptosis. This study represents the first global analysis of intracellular and exosomal miRNAs and shows that viral oncogene expression affects the abundance of multiple miRNAs likely contributing to the E6/E7-dependent growth of HPV-positive cancer cells. PMID:25760330

  15. Gene Expression Patterns of Hemizygous and Heterozygous KIT Mutations Suggest Distinct Oncogenic Pathways: A Study in NIH3T3 Cell Lines and GIST Samples

    PubMed Central

    Dessaux, Sophie; Besse, Anthony; Brahimi-Adouane, Sabrina; Emile, Jean-François; Blay, Jean-Yves; Alberti, Laurent

    2013-01-01

    Objective Most gain of function mutations of tyrosine kinase receptors in human tumours are hemizygous. Gastrointestinal stromal tumours (GIST) with homozygous mutations have a worse prognosis. We aimed to identify genes differentially regulated by hemizygous and heterozygous KIT mutations. Materials and Methods Expression of 94 genes and 384 miRNA was analysed with low density arrays in five NIH3T3 cell lines expressing the full-length human KIT cDNA wild-type (WT), hemizygous KIT mutation with del557-558 (D6) or del564-581 (D54) and heterozygous WT/D6 or WT/D54. Expression of 5 of these genes and 384 miRNA was then analysed in GISTs samples. Results Unsupervised and supervised hierarchical clustering of the mRNA and miRNA profiles showed that heterozygous mutants clustered with KIT WT expressing cells while hemizygous mutants were distinct. Among hemizygous cells, D6 and D54 expressing cells clustered separately. Most deregulated genes have been reported as potentially implicated in cancer and severals, as ANXA8 and FBN1, are highlighted by both, mRNA and miRNA analyses. MiRNA and mRNA analyses in GISTs samples confirmed that their expressions varied according to the mutation of the alleles. Interestingly, RGS16, a membrane protein of the regulator of G protein family, correlate with the subcellular localization of KIT mutants and might be responsible for regulation of the PI3K/AKT signalling pathway. Conclusion Patterns of mRNA and miRNA expression in cells and tumours depend on heterozygous/hemizygous status of KIT mutations, and deletion/presence of TYR568 & TYR570 residues. Thus each mutation of KIT may drive specific oncogenic pathways. PMID:23593401

  16. Comprehensive analysis of HPV16 integration in OSCC reveals no significant impact of physical status on viral oncogene and virally disrupted human gene expression.

    PubMed

    Olthof, Nadine C; Speel, Ernst-Jan M; Kolligs, Jutta; Haesevoets, Annick; Henfling, Mieke; Ramaekers, Frans C S; Preuss, Simon F; Drebber, Uta; Wieland, Ulrike; Silling, Steffi; Lam, Wan L; Vucic, Emily A; Kremer, Bernd; Klussmann, Jens-P; Huebbers, Christian U

    2014-01-01

    Infection with high-risk human papillomavirus (HPV) type 16 is an independent risk factor for the development of oropharyngeal squamous cell carcinomas (OSCC). However, it is unclear whether viral integration is an essential hallmark in the carcinogenic process of OSCC and whether HPV integration correlates with the level of viral gene transcription and influences the expression of disrupted host genes. We analyzed 75 patients with OSCC. HPV16-positivity was proven by p16(INK4A) immunohistochemistry, PCR and FISH. Viral integration was examined using DIPS- as well as APOT-PCR. Viral E2, E6 and E7 gene expression levels were quantified by quantitative reverse transcriptase (RT-q)PCR. Expression levels of 7 human genes disrupted by the virus were extracted from mRNA expression profiling data of 32 OSCCs. Viral copy numbers were assessed by qPCR in 73 tumors. We identified 37 HPV16-human fusion products indicating viral integration in 29 (39%) OSCC. In the remaining tumors (61%) only episome-derived PCR products were detected. When comparing OSCC with or without an integration-derived fusion product, we did not find significant differences in the mean RNA expression of viral genes E2, E6 and E7 or the viral copy numbers per cell, nor did the RNA expression of the HPV-disrupted genes differ from either group of OSCC. In conclusion, our data do not support the hypothesis that integration affects the levels of viral and/or HPV-disrupted human gene transcripts. Thus constitutive, rather than a high level, of expression of oncogene transcripts appears to be required in HPV-related OSCC. PMID:24586376

  17. Conditional overexpression of Stat3alpha in differentiating myeloid cells results in neutrophil expansion and induces a distinct, antiapoptotic and pro-oncogenic gene expression pattern.

    PubMed

    Redell, Michele S; Tsimelzon, Anna; Hilsenbeck, Susan G; Tweardy, David J

    2007-10-01

    Normal neutrophil development requires G-CSF signaling, which includes activation of Stat3. Studies of G-CSF-mediated Stat3 signaling in cell culture and transgenic mice have yielded conflicting data regarding the role of Stat3 in myelopoiesis. The specific functions of Stat3 remain unclear, in part, because two isoforms, Stat3alpha and Stat3beta, are expressed in myeloid cells. To understand the contribution of each Stat3 isoform to myelopoiesis, we conditionally overexpressed Stat3alpha or Stat3beta in the murine myeloid cell line 32Dcl3 (32D) and examined the consequences of overexpression on cell survival and differentiation. 32D cells induced to overexpress Stat3alpha, but not Stat3beta, generated a markedly higher number of neutrophils in response to G-CSF. This effect was a result of decreased apoptosis but not of increased proliferation. Comparison of gene expression profiles of G-CSF-stimulated, Stat3alpha-overexpressing 32D cells with those of cells with normal Stat3alpha expression revealed novel Stat3 gene targets, which may contribute to neutrophil expansion and improved survival, most notably Slc28a2, a purine nucleoside transporter, which is critical for maintenance of intracellular nucleotide levels and prevention of apoptosis, and Gpr65, an acid-sensing, G protein-coupled receptor with pro-oncogenic and antiapoptotic functions. PMID:17634277

  18. The DEK oncogene activates VEGF expression and promotes tumor angiogenesis and growth in HIF-1α-dependent and -independent manners.

    PubMed

    Zhang, Yanan; Liu, Jie; Wang, Shibin; Luo, Xiaoli; Li, Yang; Lv, Zhaohui; Zhu, Jie; Lin, Jing; Ding, Lihua; Ye, Qinong

    2016-04-26

    The DEK oncogene is overexpressed in various cancers and overexpression of DEK correlates with poor clinical outcome. Vascular endothelial growth factor (VEGF) is the most important regulator of tumor angiogenesis, a process essential for tumor growth and metastasis. However, whether DEK enhances tumor angiogenesis remains unclear. Here, we show that DEK is a key regulator of VEGF expression and tumor angiogenesis. Using chromatin immunoprecipitation assay, we found that DEK promoted VEGF transcription in breast cancer cells (MCF7, ZR75-1 and MDA-MB-231) by directly binding to putative DEK-responsive element (DRE) of the VEGF promoter and indirectly binding to hypoxia response element (HRE) upstream of the DRE through its interaction with the transcription factor hypoxia-inducible factor 1α (HIF-1α), a master regulator of tumor angiogenesis and growth. DEK is responsible for recruitment of HIF-1α and the histone acetyltransferase p300 to the VEGF promoter. DEK-enhanced VEGF increases vascular endothelial cell proliferation, migration and tube formation as well as angiogenesis in the chick chorioallantoic membrane. DEK promotes tumor angiogenesis and growth in nude mice in HIF-1α-dependent and -independent manners. Immunohistochemical staining showed that DEK expression positively correlates with the expression of VEGF and microvessel number in 58 breast cancer patients. Our data establish DEK as a sequence-specific binding transcription factor, a novel coactivator for HIF-1α in regulation of VEGF transcription and a novel promoter of angiogenesis. PMID:26988756

  19. A high level of liver-specific expression of oncogenic Kras(V12) drives robust liver tumorigenesis in transgenic zebrafish.

    PubMed

    Nguyen, Anh Tuan; Emelyanov, Alexander; Koh, Chor Hui Vivien; Spitsbergen, Jan M; Lam, Siew Hong; Mathavan, Sinnakaruppan; Parinov, Serguei; Gong, Zhiyuan

    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 kras(V12) specifically in the transgenic zebrafish liver. Only a high level of kras(V12) 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 kras(V12) 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 kras(V12)-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. PMID:21729876

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

    PubMed Central

    Nguyen, Anh Tuan; Emelyanov, Alexander; Koh, Chor Hui Vivien; Spitsbergen, Jan M.; Lam, Siew Hong; Mathavan, Sinnakaruppan; Parinov, Serguei; Gong, Zhiyuan

    2011-01-01

    SUMMARY 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. PMID:21729876

  1. Transcriptional expression of Epstein-Barr virus genes and proto-oncogenes in north African nasopharyngeal carcinoma.

    PubMed

    Sbih-Lammali, F; Djennaoui, D; Belaoui, H; Bouguermouh, A; Decaussin, G; Ooka, T

    1996-05-01

    Cases of nasopharyngeal carcinoma (NPC) from North Africa show an unusual bimodal age distribution. As elsewhere, the tumor is closely associated with the presence of Epstein-Barr virus (EBV). The expression of EBV genes and c-onc genes was studied in biopsy specimens from tumors at different clinical stages from 11 young (10 to 30-year-old) and 11 adult (30 to 65-year-old) patients. It was found that the two age groups do not differ in their pattern of gene expression, that there is a tendency for later stage biopsies to express more viral and c-onc transcripts, and that samples expressing larger numbers of EBV genes also tend to express many different c-onc specificities. PMID:8732865

  2. The proto-oncogene Myc drives expression of the NK cell-activating NKp30 ligand B7-H6 in tumor cells.

    PubMed

    Textor, Sonja; Bossler, Felicitas; Henrich, Kai-Oliver; Gartlgruber, Moritz; Pollmann, Julia; Fiegler, Nathalie; Arnold, Annette; Westermann, Frank; Waldburger, Nina; Breuhahn, Kai; Golfier, Sven; Witzens-Harig, Mathias; Cerwenka, Adelheid

    2016-07-01

    Natural Killer (NK) cells are innate effector cells that are able to recognize and eliminate tumor cells through engagement of their surface receptors. NKp30 is a potent activating NK cell receptor that elicits efficient NK cell-mediated target cell killing. Recently, B7-H6 was identified as tumor cell surface expressed ligand for NKp30. Enhanced B7-H6 mRNA levels are frequently detected in tumor compared to healthy tissues. To gain insight in the regulation of expression of B7-H6 in tumors, we investigated transcriptional mechanisms driving B7-H6 expression by promoter analyses. Using luciferase reporter assays and chromatin immunoprecipitation we mapped a functional binding site for Myc, a proto-oncogene overexpressed in certain tumors, in the B7-H6 promoter. Pharmacological inhibition or siRNA/shRNA-mediated knock-down of c-Myc or N-Myc significantly decreased B7-H6 expression on a variety of tumor cells including melanoma, pancreatic carcinoma and neuroblastoma cell lines. In tumor cell lines from different origin and primary tumor tissues of hepatocellular carcinoma (HCC), lymphoma and neuroblastoma, mRNA levels of c-Myc positively correlated with B7-H6 expression. Most importantly, upon inhibition or knock-down of c-Myc in tumor cells impaired NKp30-mediated degranulation of NK cells was observed. Thus, our data imply that Myc driven tumors could be targets for cancer immunotherapy exploiting the NKp30/B7-H6 axis. PMID:27622013

  3. Enforced expression of the c-myc oncogene inhibits cell differentiation by precluding entry into a distinct predifferentiation state in G/sub 0//G/sub 1/

    SciTech Connect

    Freytag, S.O.

    1988-04-01

    A broad base of data has implicated a role for the c-myc proto-oncogene in the control of the cell cycle and cell differentiation. To further define the role of myc in these processes, the authors examined the effect of enforced myc expression on several events that are thought to be important steps leading to the terminally differentiated state: (i) the ability to arrest growth in G/sub 0//G/sub 1/, (ii) the ability to replicate the genome upon initiation of the differentiation program, and (iii) the ability to loose responsiveness to mitogens and withdraw from the cell cycle. 3T3-L1 preadipocyte cell lines expressing various levels of myc mRNA were established by transfection with a recombinant myc gene under the transcriptional control of the Rous sarcoma virus (RSV) promoter. Cells that expressed high constitutive levels of pRSV myc mRNA arrested in G/sub 0//G/sub 1/ at densities similar to those of normal cells at confluence. Upon initiation of the differentiation program, such cells traversed the cell cycle with kinetics similar to those of normal cells and subsequently arrested in G/sub 0//G/sub 1/. Thus, enforced expression of myc had no effect on the ability of cells to arrest growth in G/sub 0//G/sub 1/ or to replicate the genome upon initiation of the differentiation program. Cells were then tested for their ability to reenter the cell cycle upon exposure to high concentrations of serum and for their capacity to differentiate. In contrast to normal cells, cells expressing high constitutive levels of myc RNA reentered the cell cycle when challenged with 30% serum and failed to terminally differentiate.

  4. LncRNA Khps1 Regulates Expression of the Proto-oncogene SPHK1 via Triplex-Mediated Changes in Chromatin Structure.

    PubMed

    Postepska-Igielska, Anna; Giwojna, Alena; Gasri-Plotnitsky, Lital; Schmitt, Nina; Dold, Annabelle; Ginsberg, Doron; Grummt, Ingrid

    2015-11-19

    Although thousands of long noncoding RNAs (lncRNAs) have been discovered, very little is known about their mode of action. Here we functionally characterize an E2F1-regulated lncRNA named Khps1, which is transcribed in antisense orientation to the proto-oncogene SPHK1. Khps1 activates SPHK1 expression by recruiting the histone acetyltransferase p300/CBP to the SPHK1 promoter, which leads to local changes of the chromatin structure that ensures E2F1 binding and enhances transcription. Mechanistically, this is achieved by direct association of Khps1 with a homopurine stretch upstream of the transcription start site of SPHK1, which forms a DNA-RNA triplex that anchors the lncRNA and associated effector proteins to the gene promoter. The results reveal an lncRNA- and E2F1-driven regulatory loop in which E2F1-dependent induction of antisense RNA leads to changes in chromatin structure, facilitating E2F1-dependent expression of SPHK1 and restriction of E2F1-induced apoptosis. PMID:26590717

  5. Hidden among the crowd: differential DNA methylation-expression correlations in cancer occur at important oncogenic pathways

    PubMed Central

    Mosquera Orgueira, Adrián

    2015-01-01

    DNA methylation is a frequent epigenetic mechanism that participates in transcriptional repression. Variations in DNA methylation with respect to gene expression are constant, and, for unknown reasons, some genes with highly methylated promoters are sometimes overexpressed. In this study we have analyzed the expression and methylation patterns of thousands of genes in five groups of cancer and normal tissue samples in order to determine local and genome-wide differences. We observed significant changes in global methylation-expression correlation in all the neoplasms, which suggests that differential correlation events are frequent in cancer. A focused analysis in the breast cancer cohort identified 1662 genes whose correlation varies significantly between normal and cancerous breast, but whose DNA methylation and gene expression patterns do not change substantially. These genes were enriched in cancer-related pathways and repressive chromatin features across various model cell lines, such as PRC2 binding and H3K27me3 marks. Substantial changes in methylation-expression correlation indicate that these genes are subject to epigenetic remodeling, where the differential activity of other factors break the expected relationship between both variables. Our findings suggest a complex regulatory landscape where a redistribution of local and large-scale chromatin repressive domains at differentially correlated genes (DCGs) creates epigenetic hotspots that modulate cancer-specific gene expression. PMID:26029238

  6. Tumor suppressor ASXL1 is essential for the activation of INK4B expression in response to oncogene activity and anti-proliferative signals.

    PubMed

    Wu, Xudong; Bekker-Jensen, Ida Holst; Christensen, Jesper; Rasmussen, Kasper Dindler; Sidoli, Simone; Qi, Yan; Kong, Yu; Wang, Xi; Cui, Yajuan; Xiao, Zhijian; Xu, Guogang; Williams, Kristine; Rappsilber, Juri; Sønderby, Casper Kaae; Winther, Ole; Jensen, Ole N; Helin, Kristian

    2015-11-01

    ASXL1 mutations are frequently found in hematological tumors, and loss of Asxl1 promotes myeloid transformation in mice. Here we present data supporting a role for an ASXL1-BAP1 complex in the deubiquitylation of mono-ubiquitylated lysine 119 on Histone H2A (H2AK119ub1) in vivo. The Polycomb group proteins control the expression of the INK4B-ARF-INK4A locus during normal development, in part through catalyzing mono-ubiquitylation of H2AK119. Since the activation of the locus INK4B-ARF-INK4A plays a fail-safe mechanism protecting against tumorigenesis, we investigated whether ASXL1-dependent H2A deubiquitylation plays a role in its activation. Interestingly, we found that ASXL1 is specifically required for the increased expression of p15(INK4B) in response to both oncogenic signaling and extrinsic anti-proliferative signals. Since we found that ASXL1 and BAP1 both are enriched at the INK4B locus, our results suggest that activation of the INK4B locus requires ASXL1/BAP1-mediated deubiquitylation of H2AK119ub1. Consistently, our results show that ASXL1 mutations are associated with lower expression levels of p15(INK4B) and a proliferative advantage of hematopoietic progenitors in primary bone marrow cells, and that depletion of ASXL1 in multiple cell lines results in resistance to growth inhibitory signals. Taken together, this study links ASXL1-mediated H2A deubiquitylation and transcriptional activation of INK4B expression to its tumor suppressor functions. PMID:26470845

  7. Redox-modulating agents target NOX2-dependent IKKε oncogenic kinase expression and proliferation in human breast cancer cell lines

    PubMed Central

    Mukawera, Espérance; Chartier, Stefany; Williams, Virginie; Pagano, Patrick J.; Lapointe, Réjean; Grandvaux, Nathalie

    2015-01-01

    Oxidative stress is considered a causative factor in carcinogenesis, but also in the development of resistance to current chemotherapies. The appropriate usage of redox-modulating compounds is limited by the lack of knowledge of their impact on specific molecular pathways. Increased levels of the IKKε kinase, as a result of gene amplification or aberrant expression, are observed in a substantial number of breast carcinomas. IKKε not only plays a key role in cell transformation and invasiveness, but also in the development of resistance to tamoxifen. Here, we studied the effect of in vitro treatment with the redox-modulating triphenylmethane dyes, Gentian Violet and Brilliant Green, and nitroxide Tempol on IKKε expression and cell proliferation in the human breast cancer epithelial cell lines exhibiting amplification of IKKε, MCF-7 and ZR75.1. We show that Gentian Violet, Brilliant Green and Tempol significantly decrease intracellular superoxide anion levels and inhibit IKKε expression and cell viability. Treatment with Gentian Violet and Brilliant Green was associated with a reduced cyclin D1 expression and activation of caspase 3 and/or 7. Tempol decreased cyclin D1 expression in both cell lines, while activation of caspase 7 was only observed in MCF-7 cells. Silencing of the superoxide-generating NOX2 NADPH oxidase expressed in breast cancer cells resulted in the significant reduction of IKKε expression. Taken together, our results suggest that redox-modulating compounds targeting NOX2 could present a particular therapeutic interest in combination therapy against breast carcinomas exhibiting IKKε amplification. PMID:26177467

  8. Redox-modulating agents target NOX2-dependent IKKε oncogenic kinase expression and proliferation in human breast cancer cell lines.

    PubMed

    Mukawera, Espérance; Chartier, Stefany; Williams, Virginie; Pagano, Patrick J; Lapointe, Réjean; Grandvaux, Nathalie

    2015-12-01

    Oxidative stress is considered a causative factor in carcinogenesis, but also in the development of resistance to current chemotherapies. The appropriate usage of redox-modulating compounds is limited by the lack of knowledge of their impact on specific molecular pathways. Increased levels of the IKKε kinase, as a result of gene amplification or aberrant expression, are observed in a substantial number of breast carcinomas. IKKε not only plays a key role in cell transformation and invasiveness, but also in the development of resistance to tamoxifen. Here, we studied the effect of in vitro treatment with the redox-modulating triphenylmethane dyes, Gentian Violet and Brilliant Green, and nitroxide Tempol on IKKε expression and cell proliferation in the human breast cancer epithelial cell lines exhibiting amplification of IKKε, MCF-7 and ZR75.1. We show that Gentian Violet, Brilliant Green and Tempol significantly decrease intracellular superoxide anion levels and inhibit IKKε expression and cell viability. Treatment with Gentian Violet and Brilliant Green was associated with a reduced cyclin D1 expression and activation of caspase 3 and/or 7. Tempol decreased cyclin D1 expression in both cell lines, while activation of caspase 7 was only observed in MCF-7 cells. Silencing of the superoxide-generating NOX2 NADPH oxidase expressed in breast cancer cells resulted in the significant reduction of IKKε expression. Taken together, our results suggest that redox-modulating compounds targeting NOX2 could present a particular therapeutic interest in combination therapy against breast carcinomas exhibiting IKKε amplification. PMID:26177467

  9. Proto-oncogenes II.

    PubMed

    Rosen, P

    1988-12-01

    In reviewing recent literature on activated proto-oncogenes including retroviral infection (without oncogene), translocation and inherited childhood cancer, I have come to the conclusion that activated proto-oncogenes are not involved in development of tumors. There is one exception in which a translocated proto-myc leads to transformation. That is the case of the trangenic mouse embryo where faulty development occurs. PMID:3226361

  10. Expression of the glioma-associated oncogene homolog (GLI) 1 in human breast cancer is associated with unfavourable overall survival

    PubMed Central

    2009-01-01

    Background The transcription factor GLI1, a member of the GLI subfamily of Krüppel-like zinc finger proteins is involved in signal transduction within the hedgehog pathway. Aberrant hedgehog signalling has been implicated in the development of different human tumour entities such as colon and lung cancer and increased GLI1 expression has been found in these tumour entities as well. In this study we questioned whether GLI1 expression might also be important in human breast cancer development. Furthermore we correlated GLI1 expression with histopathological and clinical data to evaluate whether GLI1 could represent a new prognostic marker in breast cancer treatment. Methods Applying semiquantitative realtime PCR analysis and immunohistochemistry (IHC) GLI1 expression was analysed in human invasive breast carcinomas (n = 229) in comparison to normal human breast tissues (n = 58). GLI1 mRNA expression was furthermore analysed in a set of normal (n = 3) and tumourous breast cell lines (n = 8). IHC data were statistically interpreted using SPSS version 14.0. Results Initial analysis of GLI1 mRNA expression in a small cohort of (n = 5) human matched normal and tumourous breast tissues showed first tendency towards GLI1 overexpression in human breast cancers. However only a small sample number was included into these analyses and values for GLI1 overexpression were statistically not significant (P = 0.251, two-tailed Mann-Whitney U-test). On protein level, nuclear GLI1 expression in breast cancer cells was clearly more abundant than in normal breast epithelial cells (P = 0.008, two-tailed Mann-Whitney U-test) and increased expression of GLI1 protein in breast tumours significantly correlated with unfavourable overall survival (P = 0.019), but also with higher tumour stage (P < 0.001) and an increased number of tumour-positive axillar lymph nodes (P = 0.027). Interestingly, a highly significant correlation was found between GLI1 expression and the expression of SHH, a

  11. Hematopoietic expression of oncogenic BRAF promotes aberrant growth of monocyte-lineage cells resistant to PLX4720

    PubMed Central

    Kamata, Tamihiro; Dankort, David; Kang, Jing; Giblett, Susan; Pritchard, Catrin A.; McMahon, Martin; Leavitt, Andrew D.

    2013-01-01

    Mutational activation of BRAF leading to expression of the BRAFV600E oncoprotein was recently identified in a high percentage of specific hematopoietic neoplasms in monocyte/histiocyte and mature B-cell lineages. Although BRAFV600E is a driver oncoprotein and pharmacological target in solid tumors such as melanoma, lung and thyroid cancer, it remains unknown whether BRAFV600E is an appropriate therapeutic target in hematopoietic neoplasms. To address this critical question, we generated a mouse model expressing inducible BRAFV600E in the hematopoietic system, and evaluated the efficacy of pathway-targeted therapeutics against primary hematopoietic cells. In this model, BRAFV600E expression conferred cytokine-independent growth to monocyte/macrophage-lineage progenitors leading to aberrant in vivo and in vitro monocyte/macrophage expansion. Furthermore, transplantation of BRAFV600E-expressing bone marrow cells promoted an in vivo pathology most notable for monocytosis in hematopoietic tissues and visceral organs. In vitro analysis revealed that MEK inhibition, but not RAF inhibition, effectively suppressed cytokine-independent clonal growth of monocyte/macrophage-lineage progenitors. However, combined RAF and PI3K inhibition effectively inhibited cytokine-independent colony formation, suggesting autocrine PI3K pathway activation. Taken together, these results provide evidence that constitutively activated BRAFV600E drives aberrant proliferation of monocyte-lineage cells. This study supports the development of pathway-targeted therapeutics in the treatment of BRAFV600E-expressing hematopoietic neoplasms in the monocyte/histiocyte lineage. PMID:24152792

  12. High Expression of the Newly Found Long Noncoding RNA Z38 Promotes Cell Proliferation and Oncogenic Activity in Breast Cancer

    PubMed Central

    Deng, Rilin; Liu, Bin; Wang, Yan; Yan, Feng; Hu, Shifan; Wang, Hongcan; Wang, Tingting; Li, Bin; Deng, Xiyun; Xiang, Shuanglin; Yang, Yinke; Zhang, Jian

    2016-01-01

    The aberrant expression of long noncoding RNAs (lncRNAs) has great impacts on cancer origination and progression. In the current study, a newly found lncRNA Z38, which was identified through combining experiments of suppression subtractive hybridization (SSH) and reverse dot-blotting, was found to have high expression in breast cancer. More importantly, inhibiting Z38 expression by gene silencing greatly suppressed breast cancer cell proliferation and tumorigenesis, and treatment with Z38 siRNAs significantly induced cell apoptosis and inhibited tumor growth. In conclusion, the newly found lncRNA Z38, which plays important roles in breast cancer, may act as a candidate biomarker and therapeutic target in carcinomas. PMID:27053956

  13. High Expression of the Newly Found Long Noncoding RNA Z38 Promotes Cell Proliferation and Oncogenic Activity in Breast Cancer.

    PubMed

    Deng, Rilin; Liu, Bin; Wang, Yan; Yan, Feng; Hu, Shifan; Wang, Hongcan; Wang, Tingting; Li, Bin; Deng, Xiyun; Xiang, Shuanglin; Yang, Yinke; Zhang, Jian

    2016-01-01

    The aberrant expression of long noncoding RNAs (lncRNAs) has great impacts on cancer origination and progression. In the current study, a newly found lncRNA Z38, which was identified through combining experiments of suppression subtractive hybridization (SSH) and reverse dot-blotting, was found to have high expression in breast cancer. More importantly, inhibiting Z38 expression by gene silencing greatly suppressed breast cancer cell proliferation and tumorigenesis, and treatment with Z38 siRNAs significantly induced cell apoptosis and inhibited tumor growth. In conclusion, the newly found lncRNA Z38, which plays important roles in breast cancer, may act as a candidate biomarker and therapeutic target in carcinomas. PMID:27053956

  14. Exposure to airborne PM2.5 suppresses microRNA expression and deregulates target oncogenes that cause neoplastic transformation in NIH3T3 cells

    PubMed Central

    Cheng, Xinxin; Shao, Mingming; Wu, Chen; Wang, Suhan; Li, Hongmin; Wei, Lixuan; Gao, Yanning; Tan, Wen; Cheng, Shujun; Wu, Tangchun; Yu, Dianke; Lin, Dongxin

    2015-01-01

    Long-term exposure to airborne PM2.5 is associated with increased lung cancer risk but the underlying mechanism remains unclear. We characterized global microRNA and mRNA expression in human bronchial epithelial cells exposed to PM2.5 organic extract and integrally analyzed microRNA-mRNA interactions. Foci formation and xenograft tumorigenesis in mice with NIH3T3 cells expressing genes targeted by microRNAs were performed to explore the oncogenic potential of these genes. We also detected plasma levels of candidate microRNAs in subjects exposed to different levels of air PM2.5 and examined the aberrant expression of genes targeted by these microRNAs in human lung cancer. Under our experimental conditions, treatment of cells with PM2.5 extract resulted in downregulation of 138 microRNAs and aberrant expression of 13 mRNAs (11 upregulation and 2 downregulation). In silico and biochemical analyses suggested SLC30A1, SERPINB2 and AKR1C1, among the upregulated genes, as target for miR-182 and miR-185, respectively. Ectopic expression of each of these genes significantly enhanced foci formation in NIH3T3 cells. Following subcutaneous injection of these cells into nude mice, fibrosarcoma were formed from SLC30A1- or SERPINB2-expressing cells. Reduced plasma levels of miR-182 were detected in subjects exposed to high level of PM2.5 than in those exposed to low level of PM2.5 (P = 0.043). Similar results were seen for miR-185 although the difference was not statistically significant (P = 0.328). Increased expressions of SLC30A1, SERPINB2 and AKR1C1 were detected in human lung cancer. These results suggest that modulation of miR-182 and miR-185 and their target genes may contribute to lung carcinogenesis attributable to PM2.5 exposure. PMID:26338969

  15. SIRT1 inhibits proliferation of pancreatic cancer cells expressing pancreatic adenocarcinoma up-regulated factor (PAUF), a novel oncogene, by suppression of {beta}-catenin

    SciTech Connect

    Cho, Il-Rae; Koh, Sang Seok; Malilas, Waraporn; Srisuttee, Ratakorn; Moon, Jeong; Choi, Young-Whan; Horio, Yoshiyuki; Oh, Sangtaek; Chung, Young-Hwa

    2012-06-29

    Highlights: Black-Right-Pointing-Pointer SIRT1 inhibits protein levels of {beta}-catenin and its transcriptional activity. Black-Right-Pointing-Pointer Nuclear localization of SIRT1 is not required for the decrease of {beta}-catenin expression. Black-Right-Pointing-Pointer SIRT1-mediated degradation of {beta}-catenin is not required for GSK-3{beta} and Siah-1 but for proteosome. Black-Right-Pointing-Pointer SIRT1 activation inhibits proliferation of pancreatic cancer cells expressing PAUF. -- Abstract: Because we found in a recent study that pancreatic adenocarcinoma up-regulated factor (PAUF), a novel oncogene, induces a rapid proliferation of pancreatic cells by up-regulation of {beta}-catenin, we postulated that {beta}-catenin might be a target molecule for pancreatic cancer treatment. We thus speculated whether SIRT1, known to target {beta}-catenin in a colon cancer model, suppresses {beta}-catenin in those pancreatic cancer cells that express PAUF (Panc-PAUF). We further evaluated whether such suppression would lead to inhibition of the proliferation of these cells. The ectopic expression of either SIRT1 or resveratrol (an activator of SIRT1) suppressed levels of {beta}-catenin protein and its transcriptional activity in Panc-PAUF cells. Conversely, suppression of SIRT1 expression by siRNA enhanced {beta}-catenin expression and transcriptional activity. SIRT1 mutant analysis showed that nuclear localization of SIRT1 is not required for reduction of {beta}-catenin. Treatment with MG132, a proteasomal inhibitor, restored {beta}-catenin protein levels, suggesting that SIRT1-mediated degradation of {beta}-catenin requires proteasomal activity. It was reported that inhibition of GSK-3{beta} or Siah-1 stabilizes {beta}-catenin in colon cancer cells, but suppression of GSK-3{beta} or Siah-1 using siRNA in the presence of resveratrol instead diminished {beta}-catenin protein levels in Panc-PAUF cells. This suggests that GSK-3{beta} and Siah-1 are not involved in SIRT1

  16. TP53: an oncogene in disguise

    PubMed Central

    Soussi, T; Wiman, K G

    2015-01-01

    The standard classification used to define the various cancer genes confines tumor protein p53 (TP53) to the role of a tumor suppressor gene. However, it is now an indisputable fact that many p53 mutants act as oncogenic proteins. This statement is based on multiple arguments including the mutation signature of the TP53 gene in human cancer, the various gains-of-function (GOFs) of the different p53 mutants and the heterogeneous phenotypes developed by knock-in mouse strains modeling several human TP53 mutations. In this review, we will shatter the classical and traditional image of tumor protein p53 (TP53) as a tumor suppressor gene by emphasizing its multiple oncogenic properties that make it a potential therapeutic target that should not be underestimated. Analysis of the data generated by the various cancer genome projects highlights the high frequency of TP53 mutations and reveals that several p53 hotspot mutants are the most common oncoprotein variants expressed in several types of tumors. The use of Muller's classical definition of mutations based on quantitative and qualitative consequences on the protein product, such as ‘amorph', ‘hypomorph', ‘hypermorph' ‘neomorph' or ‘antimorph', allows a more meaningful assessment of the consequences of cancer gene modifications, their potential clinical significance, and clearly demonstrates that the TP53 gene is an atypical cancer gene. PMID:26024390

  17. Crystallin gene expression and lentoid body formation in quail embryo neuroretina cultures transformed by the oncogenic retrovirus Mill Hill 2 or Rous sarcoma virus.

    PubMed Central

    Simonneau, L; Crisanti, P; Lorinet, A M; Alliot, F; Courtois, Y; Calothy, G; Pessac, B

    1986-01-01

    The lens-specific proteins alpha and delta crystallins and lentoid bodies, structures that follow a differentiation pathway similar to that of the lens, regularly appear after 4 to 5 weeks in quail embryo neuroretina monolayer cultures. We have investigated the effects of the avian oncogenic retroviruses Mill Hill 2 and Rous sarcoma virus on this process. Quail embryo neuroretina cells transformed by Mill Hill 2 virus were established into permanent cultures that synthesized alpha and delta crystallins and contained stem cells for the production of lentoid bodies. In contrast, transformation with the Rous sarcoma virus mutant tsNY-68 blocked the appearance of mRNA crystallins, but cytoplasmic alpha and delta crystallin mRNA and alpha crystallin appeared 44 h after a shift to the nonpermissive temperature. However, delta crystallins and lentoid bodies were only present after 7 days. The crystallins of transformed quail neuroretina cultures were immunologically indistinguishable from those of quail lenses and of normal quail embryo neuroretina cultures. Images PMID:3025609

  18. Nucleotide sequence and expression in vitro of cDNA derived from mRNA of int-1, a provirally activated mouse mammary oncogene.

    PubMed Central

    Fung, Y K; Shackleford, G M; Brown, A M; Sanders, G S; Varmus, H E

    1985-01-01

    The mouse int-1 gene is a putative mammary oncogene discovered as a target for transcriptionally activating proviral insertion mutations in mammary carcinomas induced by the mouse mammary tumor virus in C3H mice. We have isolated molecular clones of full- or nearly full-length cDNA transcribed from int-1 RNA (2.6 kilobases) in a virus-induced mammary tumor. Comparison of the nucleotide sequence of the cDNA clones with that of the int-1 gene (A. van Ooyen and R. Nusse, Cell 39:233-240, 1984) shows the following. The coding region of the int-1 gene is composed of four exons. The splice donor and acceptor sites conform to consensus; however, at least two closely spaced polyadenylation sites are used, and the transcriptional initiation site remains ambiguous. The major open reading frame is preceded by an open frame 10 codons in length. The mRNA encodes a 41-kilodalton protein with several striking features--a strongly hydrophobic amino terminus, a cysteine-rich carboxy terminus, and four potential glycosylation sites. There are no differences in nucleotide sequence between the known exons of the normal and a provirally activated allele. The length of the deduced open reading frame was further confirmed by in vitro translation of RNA transcribed from the cDNA clones with SP6 RNA polymerase. Images PMID:3018519

  19. E2F1 expression is deregulated and plays an oncogenic role in sporadic Burkitt’s lymphoma

    PubMed Central

    Molina-Privado, Irene; Rodríguez-Martínez, María; Rebollo, Patricia; Martín-Pérez, Daniel; Artiga, María-Jesús; Menárguez, Javier; Flemington, Erik K.; Piris, Miguel A.; Campanero, Miguel R.

    2009-01-01

    Current treatments of sBL are associated with severe toxicities. A better understanding of sBL formation would facilitate development of less toxic therapies. The etiology of sporadic Burkitt’s lymphoma (sBL) remains however largely unknown, being C-MYC up-regulation the only lesion known to occur in all sBL cases. Several studies examining the role of C-MYC in the pathogenesis of BL have concluded that C-MYC translocation is not the only critical event and that additional unidentified factors are expected to be involved in the formation of this tumor. We herein report that a gene distinct from C-MYC, E2F1, is involved in the formation of all or most sBL tumors. We found that E2F1 is highly expressed in Burkitt’s lymphoma cell lines and sBL lymphoma specimens. Our data indicate that its elevated expression is not merely the consequence of the presence of more cycling cells in this tumor relative to other cell lines or to other neoplasias. In fact, we show that reduction of its expression in sBL cells inhibits tumor formation and decreases their proliferation rate. We also provide data suggesting that E2F1 collaborates with C-MYC in sBL formation. E2F1 expression down-regulation did not affect, however, proliferation of human primary diploid fibroblasts. Since E2F1 is not needed for cell proliferation of normal cells, our results reveal E2F1 as a promising therapeutic target for sBL. PMID:19406837

  20. Imaging Transgene Expression with Radionuclide Imaging Technologies1

    PubMed Central

    Gambhir, SS; Herschman, HR; Cherry, SR; Barrio, JR; Satyamurthy, N; Toyokuni, T; Phelps, ME; Larson, SM; Balaton, J; Finn, R; Sadelain, M; Tjuvajev, J

    2000-01-01

    Abstract A variety of imaging technologies are being investigated as tools for studying gene expression in living subjects. Noninvasive, repetitive and quantitative imaging of gene expression will help both to facilitate human gene therapy trials and to allow for the study of animal models of molecular and cellular therapy. Radionuclide approaches using single photon emission computed tomography (SPECT) and positron emission tomography (PET) are the most mature of the current imaging technologies and offer many advantages for imaging gene expression compared to optical and magnetic resonance imaging (MRI)-based approaches. These advantages include relatively high sensitivity, full quantitative capability (for PET), and the ability to extend small animal assays directly into clinical human applications. We describe a PET scanner (micro PET) designed specifically for studies of small animals. We review “marker/reporter gene” imaging approaches using the herpes simplex type 1 virus thymidine kinase (HSV1-tk) and the dopamine type 2 receptor (D2R) genes. We describe and contrast several radiolabeled probes that can be used with the HSV1-tk reporter gene both for SPECT and for PET imaging. We also describe the advantages/disadvantages of each of the assays developed and discuss future animal and human applications. PMID:10933072

  1. E2F-Rb Complexes Assemble and Inhibit cdc25A Transcription in Cervical Carcinoma Cells following Repression of Human Papillomavirus Oncogene Expression

    PubMed Central

    Wu, Lingling; Goodwin, Edward C.; Naeger, Lisa Kay; Vigo, Elena; Galaktionov, Konstantin; Helin, Kristian; DiMaio, Daniel

    2000-01-01

    Expression of the bovine papillomavirus E2 protein in cervical carcinoma cells represses expression of integrated human papillomavirus (HPV) E6/E7 oncogenes, followed by repression of the cdc25A gene and other cellular genes required for cell cycle progression, resulting in dramatic growth arrest. To explore the mechanism of repression of cell cycle genes in cervical carcinoma cells following E6/E7 repression, we analyzed regulation of the cdc25A promoter, which contains two consensus E2F binding sites and a consensus E2 binding site. The wild-type E2 protein inhibited expression of a luciferase gene linked to the cdc25A promoter in HT-3 cervical carcinoma cells. Mutation of the distal E2F binding site in the cdc25A promoter abolished E2-induced repression, whereas mutation of the proximal E2F site or the E2 site had no effect. None of these mutations affected the activity of the promoter in the absence of E2 expression. Expression of the E2 protein also led to posttranscriptional increase in the level of E2F4, p105Rb, and p130 and induced the formation of nuclear E2F4-p130 and E2F4-p105Rb complexes. This resulted in marked rearrangement of the protein complexes that formed at the distal E2F site in the cdc25A promoter, including the replacement of free E2F complexes with E2F4-p105Rb complexes. These experiments indicated that repression of E2F-responsive promoters following HPV E6/E7 repression was mediated by activation of the Rb tumor suppressor pathway and the assembly of repressing E2F4-Rb DNA binding complexes. Importantly, these experiments revealed that HPV-induced alterations in E2F transcription complexes that occur during cervical carcinogenesis are reversed by repression of HPV E6/E7 expression. PMID:10982822

  2. The Dioxin Receptor Regulates the Constitutive Expression of the Vav3 Proto-Oncogene and Modulates Cell Shape and Adhesion

    PubMed Central

    Carvajal-Gonzalez, Jose M.; Mulero-Navarro, Sonia; Roman, Angel Carlos; Sauzeau, Vincent; Merino, Jaime M.; Bustelo, Xose R.

    2009-01-01

    The dioxin receptor (AhR) modulates cell plasticity and migration, although the signaling involved remains unknown. Here, we report a mechanism that integrates AhR into these cytoskeleton-related functions. Immortalized and mouse embryonic fibroblasts lacking AhR (AhR−/−) had increased cell area due to spread cytoplasms that reverted to wild-type morphology upon AhR re-expression. The AhR-null phenotype included increased F-actin stress fibers, depolarized focal adhesions, and enhanced spreading and adhesion. The cytoskeleton alterations of AhR−/− cells were due to down-regulation of constitutive Vav3 expression, a guanosine diphosphate/guanosine triphosphate exchange factor for Rho/Rac GTPases and a novel transcriptional target of AhR. AhR was recruited to the vav3 promoter and maintained constitutive mRNA expression in a ligand-independent manner. Consistently, AhR−/− fibroblasts had reduced Rac1 activity and increased activation of the RhoA/Rho kinase (Rock) pathway. Pharmacological inhibition of Rac1 shifted AhR+/+ fibroblasts to the null phenotype, whereas Rock inhibition changed AhR-null cells to the AhR+/+ morphology. Knockdown of vav3 transcripts by small interfering RNA induced cytoskeleton defects and changes in adhesion and spreading mimicking those of AhR-null cells. Moreover, vav3−/− MEFs, as AhR−/− mouse embryonic fibroblasts, had increased cell area and enhanced stress fibers. By modulating Vav3-dependent signaling, AhR could regulate cell shape, adhesion, and migration under physiological conditions and, perhaps, in certain pathological states. PMID:19158396

  3. Activating the Expression of Human K-rasG12D Stimulates Oncogenic Transformation in Transgenic Goat Fetal Fibroblast Cells

    PubMed Central

    Gong, Jianhua; Wang, Zhongde; Polejaeva, Irina; Salgia, Ravi; Kao, Chien-Min; Chen, Chin-Tu; Chen, Guangchun; Chen, Liaohai

    2014-01-01

    Humane use of preclinical large animal cancer models plays a critical role in understanding cancer biology and developing therapeutic treatments. Among the large animal candidates, goats have great potentials as sustainable sources for large animal cancer model development. Goats are easier to handle and cheaper to raise. The genome of the goats has been sequenced recently. It has been known that goats develop skin, adrenal cortex, breast and other types of cancers. Technically, goats are subject to somatic cell nuclear transfer more efficiently and exhibit better viability through the cloning process. Towards the development of a goat cancer model, we created a transgenic goat fetal fibroblast (GFF) cell as the donor cell for SCNT. Human mutated K-ras (hK-rasG12D) was chosen as the transgene, as it is present in 20% of cancers. Both hK-rasG12D and a herpes simplex viral thymidine kinase (HSV1-tk) reporter genes, flanked by a pair of LoxP sites, were knocked in the GFF endogenous K-ras locus through homologous recombination. Following Cre-mediated activation (with a 95% activation efficiency), hK-rasG12D and HSV1-tk were expressed in the transgenic GFF cells, evidently through the presence of corresponding mRNAs, and confirmed by HSV1-tk protein function assay. The hK-rasG12D expressing GFF cells exhibited enhanced proliferation rates and an anchorage-independent growth behavior. They were able to initiate tumor growth in athymic nude mice. In conclusion, after activating hK-rasG12D gene expression, hK-rasG12D transgenic GFF cells were transformed into tumorgenesis cells. Transgenic goats via SCNT using the above-motioned cells as the donor cells have been established. PMID:24594684

  4. Oncogenic NanogP8 expression regulates cell proliferation and migration through the Akt/mTOR signaling pathway in human gastric cancer – SGC-7901cell line

    PubMed Central

    Jiang, Zheng; Liu, Yao; Wang, Chuan

    2016-01-01

    Background Although elevated expression of NanogP8 has been detected in many human tumor tissues, its role in gastric tumorigenesis remains unclear. Therefore, this study aimed to investigate the function and regulatory mechanism of NanogP8 in gastric cancer. Methods In this study, NanogP8 cDNA was amplified by real time polymerase chain reaction from the human gastric cancer cell line SGC-7901. The shRNA for RNA interference was established. The NanogP8, pAkt, Akt, pERK, ERK, p-mTOR, and mTOR proteins were detected by using the Western blot assay. Cell viability was evaluated by using the CCK-8 assay. Cell migration and invasion were also examined by using the transwell assay. Results The results indicated that the NanogP8 overexpression promoted proliferation and migration of SGC-7901 cell line, whereas its ablation exerted opposite effects. Interestingly, NanogP8 activated Akt, a key mediator of survival signals, and without affecting total Akt protein level. The NanogP8-increased gastric cell proliferation was downregulated by Akt inhibition. Our results further showed that increasing NanogP8 expression in human gastric cancer cells promoted cell proliferation by activating the AKT/mTOR pathway and further maintained gastric cell survival. Conclusion Our findings extend the knowledge regarding the oncogenic functions and proved that the NanogP8 regulates cell proliferation and migration by Akt/mTOR signaling pathway in human gastric cancer SGC-7901cell line. PMID:27563247

  5. Combining High-Content Imaging and Phenotypic Classification Analysis of Senescence-Associated Beta-Galactosidase Staining to Identify Regulators of Oncogene-Induced Senescence.

    PubMed

    Chan, Keefe T; Paavolainen, Lassi; Hannan, Katherine M; George, Amee J; Hannan, Ross D; Simpson, Kaylene J; Horvath, Peter; Pearson, Richard B

    2016-09-01

    Hyperactivation of the PI3K/AKT/mTORC1 signaling pathway is a hallmark of the majority of sporadic human cancers. Paradoxically, chronic activation of this pathway in nontransformed cells promotes senescence, which acts as a significant barrier to malignant progression. Understanding how this oncogene-induced senescence is maintained in nontransformed cells and conversely how it is subverted in cancer cells will provide insight into cancer development and potentially identify novel therapeutic targets. High-throughput screening provides a powerful platform for target discovery. Here, we describe an approach to use RNAi transfection of a pre-established AKT-induced senescent cell population and subsequent high-content imaging to screen for senescence regulators. We have incorporated multiparametric readouts, including cell number, proliferation, and senescence-associated beta-galactosidase (SA-βGal) staining. Using machine learning and automated image analysis, we also describe methods to classify distinct phenotypes of cells with SA-βGal staining. These methods can be readily adaptable to high-throughput functional screens interrogating the mechanisms that maintain and prevent senescence in various contexts. PMID:27552145

  6. The rs391957 variant cis-regulating oncogene GRP78 expression contributes to the risk of hepatocellular carcinoma.

    PubMed

    Zhu, Xiao; Zhang, Jinfang; Fan, Wenguo; Wang, Fang; Yao, Hong; Wang, Zifeng; Hou, Shengping; Tian, Yinghong; Fu, Weiming; Xie, Dan; Zhu, Wei; Long, Jun; Wu, Leijie; Zheng, Xuebao; Kung, Hsiangfu; Zhou, Keyuan; Lin, Marie C M; Luo, Hui; Li, Dongpei

    2013-06-01

    Glucose-regulated protein 78 (GRP78) is one of the most important responders to disease-related stress. We assessed the association of the promoter polymorphisms of GRP78 with risk of hepatocellular carcinoma (HCC) and GRP78 expression in a Chinese population. We examined 1007 patients undergoing diagnostic HCC and 810 unrelated healthy controls. Mechanisms by which the GRP78 promoter polymorphism modulates HCC risk and GRP78 levels were analyzed. The promoter haplotype and diplotype carrying rs391957 (-415bp) allele G and genotype GG was strongly associated with HCC risk. Luciferase reporter assays indicated that the promoter carrying rs391957 allele G (haplotype GCCd) showed increased activity in HepG2 cells and Hela cells. rs391957 was also shown to increase the affinity of the transcriptional activator Ets-2, the resistance to apoptosis, as well as cell instability in stressful microenvironment. Furthermore, compared with allele A, rs391957 allele G was associated with higher levels of GRP78 mRNA and protein in HCC tissues. These findings provided new insights into the pathogenesis of HCC and an unexpected effect of the interaction between rs391957 and Ets-2 on hepatocarcinogenesis, and especially supported the hypothesis that stress-related and evolutionarily conserved genetic variant(s) influencing transcriptional regulation could predict susceptibilities. PMID:23416888

  7. Suppressor of cytokine signaling 1-dependent regulation of the expression and oncogenic functions of p21(CIP1/WAF1) in the liver.

    PubMed

    Yeganeh, M; Gui, Y; Kandhi, R; Bobbala, D; Tobelaim, W-S; Saucier, C; Yoshimura, A; Ferbeyre, G; Ramanathan, S; Ilangumaran, S

    2016-08-11

    tumor-suppressor functions of SOCS1 in the liver could be mediated, at least partly, via regulation of the expression, stability and subcellular distribution of p21 and its paradoxical oncogenic functions, namely, resistance to apoptosis and increased proliferation. PMID:26725321

  8. Inhibition of MEK5 by BIX02188 induces apoptosis in cells expressing the oncogenic mutant FLT3-ITD

    SciTech Connect

    Razumovskaya, Elena; Sun, Jianmin; Roennstrand, Lars

    2011-08-26

    Highlights: {yields} In this study we have demonstrated that FLT3 activation leads to activation of ERK5. {yields} We have demonstrated that ERK5 is involved in activation of AKT downstream of FLT3. {yields} (BIX02188) blocks activation of ERK5 and induces apoptosis in FLT3 Ba/F3 cells. {yields} (BIX02188) induce apoptosis in the two leukemic cell lines MV4-11 and MOLM-13. -- Abstract: Fms-like tyrosine kinase-3 (FLT3) is a growth factor receptor normally expressed on hematopoietic progenitor cells. Approximately one third of all patients with AML carry an activating mutation in FLT3 that drives proliferation and survival of the leukemic cells. The most common activating mutation is the so-called internal tandem duplication (ITD), which involves an in-frame duplication of a segment of varying length in the region of the FLT3 gene that encodes the juxtamembrane domain. The pathways downstream of FLT3-ITD are partially known but further knowledge regarding the downstream signal transduction molecules is important in order to develop alternative strategies for pharmacological intervention. In this paper we have studied the role of MEK/ERK5 in FLT3-ITD mediated transformation. We have found that both wild-type FLT3 and FLT3-ITD activate MEK5 leading to the activation of ERK5. By use of the selective inhibitor of MEK5, (BIX02188), we have shown that activation of AKT downstream of FLT3 is partially dependent on ERK5. Furthermore, inhibition of MEK5/ERK5 induces apoptosis of both FLT3-ITD transfected Ba/F3 cells as well as the FLT3-ITD carrying leukemic cell lines MV4-11 and MOLM-13. These results suggest that MEK5/ERK5 is important for FLT3-ITD induced hematopoietic transformation and may thus represent an alternative therapeutic target in the treatment of FLT3-ITD positive leukemia.

  9. Live Imaging and Gene Expression Analysis in Zebrafish Identifies a Link between Neutrophils and Epithelial to Mesenchymal Transition

    PubMed Central

    Freisinger, Christina M.; Huttenlocher, Anna

    2014-01-01

    Chronic inflammation is associated with epithelial to mesenchymal transition (EMT) and cancer progression however the relationship between inflammation and EMT remains unclear. Here, we have exploited zebrafish to visualize and quantify the earliest events during epithelial cell transformation induced by oncogenic HRasV12. Live imaging revealed that expression of HRasV12 in the epidermis results in EMT and chronic neutrophil and macrophage infiltration. We have developed an in vivo system to probe and quantify gene expression changes specifically in transformed cells from chimeric zebrafish expressing oncogenic HRasV12 using translating ribosomal affinity purification (TRAP). We found that the expression of genes associated with EMT, including slug, vimentin and mmp9, are enriched in HRasV12 transformed epithelial cells and that this enrichment requires the presence of neutrophils. An early signal induced by HRasV12 in epithelial cells is the expression of il-8 (cxcl8) and we found that the chemokine receptor, Cxcr2, mediates neutrophil but not macrophage recruitment to the transformed cells. Surprisingly, we also found a cell autonomous role for Cxcr2 signaling in transformed cells for both neutrophil recruitment and EMT related gene expression associated with Ras transformation. Taken together, these findings implicate both autocrine and paracrine signaling through Cxcr2 in the regulation of inflammation and gene expression in transformed epithelial cells. PMID:25372289

  10. Diagnostic correlation between RET proto-oncogene mutation, imaging techniques, biochemical markers and morphological examination in MEN2A syndrome: case report and literature review.

    PubMed

    Sovrea, Alina Simona; Dronca, Eleonora; Galatâr, Mihaela; Radian, Serban; Vornicescu, Corina; Georgescu, Carmen

    2014-01-01

    Multiple endocrine neoplasia type 2 (MEN2) is a rare autosomal dominant monogenic disorder caused mostly by missense mutations in the RET (REarranged during Transfection) proto-oncogene on chromosome 10q11.2. MEN2A represents more than 50% of all MEN2 cases, having a regular pattern with medullary thyroid carcinoma (MTC) incidence of 90-100%, bilateral pheochromocytoma (PCC) incidence of 40-50% and primary hyperparathyroidism (HPT) incidence of 10-25%. Until recently, the diagnosis of MTC was most frequently based on fine-needle aspiration of thyroid nodules, after an ultrasound examination and endocrine evaluation of serum calcitonin levels. Nowadays, RET gene screening (starting with exons 10 and 11) is a mandatory test used for identification of both symptomatic and non-symptomatic MTC carriers or for exclusion of healthy individuals from subsequent periodical clinical/biochemical screening. In this context, and in the idea of PCC preceding MTC, the early detection of germline RET mutations are highly suggestive for hereditary disease. PCC diagnosis is established in classical manner by abdominal ultrasound imaging or computed tomography confirming the presence of adrenal gland masses, elevated plasma metanephrines and normetanephrines values and histopathological examination. Additional HPT diagnosis is acknowledged by serum ionized calcium and parathormone levels. Here we report a hereditary case of MEN2A in a two-generation Romanian family, along with data presenting the importance of correlative plurifactorial diagnostic scheme in this syndrome and a short literature review. PMID:24969991

  11. Molecular imaging of in vivo gene expression

    PubMed Central

    Harney, Allison S.; Meade, Thomas J.

    2015-01-01

    Background Advances in imaging technologies have taken a prominent role in experimental and translational research and provide essential information on how changes in gene expression are related to downstream developmental and disease states. Discussion Magnetic resonance imaging contrast agents and optical probes developed to enhance signal intensity in the presence of a specific enzyme, genetic marker, second messenger or metabolite can prove a facile method of advancing the understanding of molecular events in disease progression. Conclusion The ability to detect changes in gene expression at the early stages of disease will lead to a greater understanding of disease progression, the use of early therapeutic intervention to increase patient survival, and tailored therapies to the detected genetic alterations in individual patients. PMID:21426178

  12. Imaging gene expression in single living cells

    PubMed Central

    Shav-Tal, Yaron; Singer, Robert H.; Darzacq, Xavier

    2016-01-01

    Technical advances in the field of live-cell imaging have introduced the cell biologist to a new, dynamic, subcellular world. The static world of molecules in fixed cells has now been extended to the time dimension. This allows the visualization and quantification of gene expression and intracellular trafficking events of the studied molecules and the associated enzymatic processes in individual cells, in real time. PMID:15459666

  13. The Effect of Lactobacillus crispatus and Lactobacillus rhamnosusCulture Supernatants on Expression of Autophagy Genes and HPV E6 and E7 Oncogenes in The HeLa Cell Line

    PubMed Central

    Motevaseli, Elahe; Azam, Rosa; Akrami, Seyed Mohammad; Mazlomy, Mohammadali; Saffari, Mojtaba; Modarressi, Mohammad Hossein; Daneshvar, Maryam; Ghafouri-Fard, Soudeh

    2016-01-01

    Objective The aim of this study was to clarify the mechanism by which lactobacilli exert their cytotoxic effects on cervical cancer cells. In addition, we aimed to evalu- ate the effect of lactobacilli on the expression of human papilloma virus (HPV) onco- genes. Materials and Methods In this experimental study, using quantitative real-time polymer- ase chain reaction (PCR), we analyzed the expression of CASP3 and three autophagy genes [ATG14, BECN1 and alpha 2 catalytic subunit of AMPK (PRKAA2)] along with HPV18 E6 and E7 genes in HeLa cells before and after treatment with Lactobacillus crispatus and Lactobacillus rhamnosus culture supernatants. Results The expression of CASP3 and autophagy genes in HeLa cells was de- creased after treatment with lactobacilli culture supernatants. However, this de- crease was not significant for PRKAA2 when compared with controls. In addition, expression of HPV E6 was significantly decreased after treatment with lactobacilli culture supernatants. Conclusion Lactobacilli culture supernatants can decrease expression of ATG14 and BECN1 as well as the HPV E6 oncogene. It has been demonstrated that the main changes occurring during cervical carcinogenesis in cell machinery can be reversed by suppression of HPV oncogenes. Therefore, downregulation of HPV E6 by lacto- bacilli may have therapeutic potential for cervical cancer. As the role of autophagy in cancer is complicated, further work is required to clarify the link between downregula- tion of autophagy genes and antiproliferative effects exerted by lactobacilli. PMID:26862519

  14. Inhibition of Prostaglandin Reductase 2, a Putative Oncogene Overexpressed in Human Pancreatic Adenocarcinoma, Induces Oxidative Stress-Mediated Cell Death Involving xCT and CTH Gene Expressions through 15-Keto-PGE2

    PubMed Central

    Chang, Emily Yun-Chia; Chang, Yi-Cheng; Shun, Chia-Tung; Tien, Yu-Wen; Tsai, Shu-Huei; Hee, Siow-Wey; Chen, Ing-Jung; Chuang, Lee-Ming

    2016-01-01

    Prostaglandin reductase 2 (PTGR2) is the enzyme that catalyzes 15-keto-PGE2, an endogenous PPARγ ligand, into 13,14-dihydro-15-keto-PGE2. Previously, we have reported a novel oncogenic role of PTGR2 in gastric cancer, where PTGR2 was discovered to modulate ROS-mediated cell death and tumor transformation. In the present study, we demonstrated the oncogenic potency of PTGR2 in pancreatic cancer. First, we observed that the majority of the human pancreatic ductal adenocarcinoma tissues was stained positive for PTGR2 expression but not in the adjacent normal parts. In vitro analyses showed that silencing of PTGR2 expression enhanced ROS production, suppressed pancreatic cell proliferation, and promoted cell death through increasing 15-keto-PGE2. Mechanistically, silencing of PTGR2 or addition of 15-keto-PGE2 suppressed the expressions of solute carrier family 7 member 11 (xCT) and cystathionine gamma-lyase (CTH), two important providers of intracellular cysteine for the generation of glutathione (GSH), which is widely accepted as the first-line antioxidative defense. The oxidative stress-mediated cell death after silencing of PTGR2 or addition of 15-keto-PGE2 was further abolished after restoring intracellular GSH concentrations and cysteine supply by N-acetyl-L-cysteine and 2-Mercaptomethanol. Our data highlight the therapeutic potential of targeting PTGR2/15-keto-PGE2 for pancreatic cancer. PMID:26820738

  15. Inhibition of Prostaglandin Reductase 2, a Putative Oncogene Overexpressed in Human Pancreatic Adenocarcinoma, Induces Oxidative Stress-Mediated Cell Death Involving xCT and CTH Gene Expressions through 15-Keto-PGE2.

    PubMed

    Chang, Emily Yun-Chia; Chang, Yi-Cheng; Shun, Chia-Tung; Tien, Yu-Wen; Tsai, Shu-Huei; Hee, Siow-Wey; Chen, Ing-Jung; Chuang, Lee-Ming

    2016-01-01

    Prostaglandin reductase 2 (PTGR2) is the enzyme that catalyzes 15-keto-PGE2, an endogenous PPARγ ligand, into 13,14-dihydro-15-keto-PGE2. Previously, we have reported a novel oncogenic role of PTGR2 in gastric cancer, where PTGR2 was discovered to modulate ROS-mediated cell death and tumor transformation. In the present study, we demonstrated the oncogenic potency of PTGR2 in pancreatic cancer. First, we observed that the majority of the human pancreatic ductal adenocarcinoma tissues was stained positive for PTGR2 expression but not in the adjacent normal parts. In vitro analyses showed that silencing of PTGR2 expression enhanced ROS production, suppressed pancreatic cell proliferation, and promoted cell death through increasing 15-keto-PGE2. Mechanistically, silencing of PTGR2 or addition of 15-keto-PGE2 suppressed the expressions of solute carrier family 7 member 11 (xCT) and cystathionine gamma-lyase (CTH), two important providers of intracellular cysteine for the generation of glutathione (GSH), which is widely accepted as the first-line antioxidative defense. The oxidative stress-mediated cell death after silencing of PTGR2 or addition of 15-keto-PGE2 was further abolished after restoring intracellular GSH concentrations and cysteine supply by N-acetyl-L-cysteine and 2-Mercaptomethanol. Our data highlight the therapeutic potential of targeting PTGR2/15-keto-PGE2 for pancreatic cancer. PMID:26820738

  16. The RET oncogene in papillary thyroid carcinoma.

    PubMed

    Prescott, Jason D; Zeiger, Martha A

    2015-07-01

    Papillary thyroid carcinoma (PTC) is the most common form of thyroid cancer, accounting for greater than 80% of cases. Surgical resection, with or without postoperative radioiodine therapy, remains the standard of care for patients with PTC, and the prognosis is generally excellent with appropriate treatment. Despite this, significant numbers of patients will not respond to maximal surgical and medical therapy and ultimately will die from the disease. This mortality reflects an incomplete understanding of the oncogenic mechanisms that initiate, drive, and promote PTC. Nonetheless, significant insights into the pathologic subcellular events underlying PTC have been discovered over the last 2 decades, and this remains an area of significant research interest. Chromosomal rearrangements resulting in the expression of fusion proteins that involve the rearranged during transfection (RET) proto-oncogene were the first oncogenic events to be identified in PTC. Members of this fusion protein family (the RET/PTC family) appear to play an oncogenic role in approximately 20% of PTCs. Herein, the authors review the current understanding of the clinicopathologic role of RET/PTC fusion proteins in PTC development and progression and the molecular mechanisms by which RET/PTCs exert their oncogenic effects on the thyroid epithelium. PMID:25731779

  17. Targeted expression of the E6 and E7 oncogenes of human papillomavirus type 16 in the epidermis of transgenic mice elicits generalized epidermal hyperplasia involving autocrine factors.

    PubMed Central

    Auewarakul, P; Gissmann, L; Cid-Arregui, A

    1994-01-01

    The E6 and E7 early genes of human papillomavirus type 16 have been shown in vitro to play a central role in the transforming capability of this virus. To explore their effects on differentiating epithelial cells in vivo, we used a bovine cytokeratin 10 (K10) promoter to target the expression of E6 and E7 to the suprabasal layers of the epidermis of transgenic mice. In two different lines of mice efficiently expressing the transgene, animals displayed generalized epidermal hyperplasia, hyperkeratosis and parakeratosis in the skin and the forestomach, both known to be sites of K10 expression. Northern (RNA) blot analysis revealed high levels of E6 and E7 transcripts, and in situ hybridizations localized these transcripts to the suprabasal strata of epidermis. In vivo labeling of proliferating cells showed two distinct effects of E6 and E7 expression in the epidermis: (i) an increase in the number of growing cells in the undifferentiated basal layer and (ii) abnormal proliferation of differentiated cells in the suprabasal strata. The expression of c-myc in the skin of transgenics was higher than that in control animals. The induction of c-myc transcription by topical application of tetradecanoyl phorbol acetate was prevented by simultaneous treatment with transforming growth factor beta 1 in nontransgenic skin but not in transgenic skin. In addition, transforming growth factor alpha was found to be overexpressed in the suprabasal layers of the transgenic epidermis. These findings suggest that autocrine mechanisms are involved in the development and maintenance of epidermal hyperplasia. Animals of both lines developed papillomas in skin sites exposed to mechanical irritation and wounding, suggesting that secondary events are necessary for progression to neoplasia. Collectively, these results provide new insights into the tumor promoter activities of human papillomavirus type 16 in epithelial cells in vivo. Images PMID:7969162

  18. The human oncogenic viruses

    SciTech Connect

    Luderer, A.A.; Weetall, H.H

    1986-01-01

    This book contains eight selections. The titles are: Cytogenetics of the Leukemias and Lymphomas; Cytogenetics of Solid Tumors: Renal Cell Carcinoma, Malignant Melanoma, Retinoblastoma, and Wilms' Tumor; Elucidation of a Normal Function for a Human Proto-Oncogene; Detection of HSV-2 Genes and Gene Products in Cervical Neoplasia; Papillomaviruses in Anogennital Neoplasms; Human Epstein-Barr Virus and Cancer; Hepatitis B Virus and Hepatocellular Carcinoma; and Kaposi's Sarcoma: Acquired Immunodeficiency Syndrome (AIDS) and Associated Viruses.

  19. [Hypophosphatemic oncogenic osteomalacia].

    PubMed

    Mátyus, J; Szebenyi, B; Rédl, P; Mikita, J; Gáspár, L; Haris, A; Radó, J; Kakuk, G

    2000-12-17

    The first case of oncogen osteomalacia in Hungary is reported, to draw the attention of the medical profession to it and to present the new data about its pathomechanism. Pathological hip fracture caused by hypophosphataemic osteomalacia due to isolated renal phosphate wasting was found in a previously healthy 19 years old sportsman. In spite of daily 1.5 micrograms calcitriol treatment and phosphate supplementation, hypophosphataemia persisted for 13 years and he needed regular indometacin medication for his bone pain. During that time an 1.5 cm gingival tumour was found and radically removed. The serum phosphate level returned to normal in a few hours after the operation (preoperative 0.51, after 2, 4 and 8 hours 0.61, 0.68 and 0.79 mmol/l respectively), and remained normal without calcitriol. The histological examination showed epulis with fibroblast and vascular cell proliferation, which has never been previously reported in connection with oncogenic osteomalacia. The pain resolved after 3 months and the bone density became normal in one year. Oncogenic osteomalacia must be considered in every case presenting with atypical hypophosphataemic osteomalacia. Careful dental examination is needed also in the course of search for the underlying tumour. Every tumour-like growth, even the common epulis, has to be operated radically and serum phosphate monitored in the postoperative period in all such cases. PMID:11196239

  20. Oncogenic Activities of Human Papillomaviruses

    PubMed Central

    McLaughlin-Drubin, Margaret E.; Münger, Karl

    2009-01-01

    Infectious etiologies for certain human cancers have long been suggested by epidemiological studies and studies with animals. Important support for this concept came from the discovery by Harald zur Hausen’s group that human cervical carcinoma almost universally contains certain “high-risk” human papillomavirus (HPV) types. Over the years, much has been learned about the carcinogenic activities of high-risk HPVs. These studies have revealed that two viral proteins, E6 and E7, that are consistently expressed in HPV-associated carcinomas, are necessary for induction and maintenance of the transformed phenotype. Hence, HPV-associated tumors are unique amongst human solid tumors in that they are universally caused by exposure to the same, molecularly defined oncogenic agents, and the molecular signal transduction pathways subverted by these viral transforming agents are frequently disrupted in other, non-virus associated human cancers. PMID:19540281

  1. Avian myeloblastosis virus and E26 virus oncogene products are nuclear proteins.

    PubMed Central

    Boyle, W J; Lampert, M A; Lipsick, J S; Baluda, M A

    1984-01-01

    The defective acute leukemia viruses avian myeloblastosis virus (AMV) and E26 virus each contain an inserted cellular sequence related to the same highly conserved cellular gene, proto-amv. The oncogenes of these two retroviruses differ from this cellular proto-oncogene in gene structure, transcript structure, and gene product. The product of the AMV oncogene (myb) is a 48,000 Mr protein, p48myb, encoded by a transduced segment (amv) of proto-amv flanked by short helper-virus-derived terminal sequences. The E26 virus oncogene product is a 135,000 Mr protein, p135gag-amve-ets, encoded by significant portions of a viral structural gene (gag), sequences related to proto-amv (amve), and additional E26-specific sequences (ets) transduced from cellular proto-ets. Both p48myb and p135gag-amve-ets transforming proteins are located in the nucleus of cells transformed by these viruses. A protein of 110,000 Mr which is specifically immunoprecipitated by antisera to amv peptides and may be the product of the normal cellular gene (proto-amv) has been located in the cytoplasm of cells that express proto-amv mRNA. Images PMID:6087315

  2. MYC oncogene overexpression drives renal cell carcinoma in a mouse model through glutamine metabolism

    PubMed Central

    Shroff, Emelyn H.; Eberlin, Livia S.; Dang, Vanessa M.; Gouw, Arvin M.; Gabay, Meital; Adam, Stacey J.; Bellovin, David I.; Tran, Phuoc T.; Philbrick, William M.; Garcia-Ocana, Adolfo; Casey, Stephanie C.; Li, Yulin; Dang, Chi V.; Zare, Richard N.; Felsher, Dean W.

    2015-01-01

    The MYC oncogene is frequently mutated and overexpressed in human renal cell carcinoma (RCC). However, there have been no studies on the causative role of MYC or any other oncogene in the initiation or maintenance of kidney tumorigenesis. Here, we show through a conditional transgenic mouse model that the MYC oncogene, but not the RAS oncogene, initiates and maintains RCC. Desorption electrospray ionization–mass-spectrometric imaging was used to obtain chemical maps of metabolites and lipids in the mouse RCC samples. Gene expression analysis revealed that the mouse tumors mimicked human RCC. The data suggested that MYC-induced RCC up-regulated the glutaminolytic pathway instead of the glycolytic pathway. The pharmacologic inhibition of glutamine metabolism with bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl)ethyl sulfide impeded MYC-mediated RCC tumor progression. Our studies demonstrate that MYC overexpression causes RCC and points to the inhibition of glutamine metabolism as a potential therapeutic approach for the treatment of this disease. PMID:25964345

  3. Prospective on the potential of imaging gene expression

    SciTech Connect

    Taylor, Scott E; Budinger, Thomas F.

    2000-06-01

    The feasibility of the non-invasive imaging of gene expression is explored. Calculations of the possibility of the direct imaging of specific messenger RNA with radiolabeled antisense are discussed. In addition, possible mechanism for the amplification of the biological signal to enhance image detection are discussed.

  4. Silent assassin: oncogenic ras directs epigenetic inactivation of target genes.

    PubMed

    Cheng, Xiaodong

    2008-01-01

    Oncogenic transformation is associated with genetic changes and epigenetic alterations. A study now shows that oncogenic Ras uses a complex and elaborate epigenetic silencing program to specifically repress the expression of multiple unrelated cancer-suppressing genes through a common pathway. These results suggest that cancer-related epigenetic modifications may arise through a specific and instructive mechanism and that genetic changes and epigenetic alterations are intimately connected and contribute to tumorigenesis cooperatively. PMID:18385037

  5. Oncogenic potential of guanine nucleotide stimulatory factor alpha subunit in thyroid glands of transgenic mice.

    PubMed Central

    Michiels, F M; Caillou, B; Talbot, M; Dessarps-Freichey, F; Maunoury, M T; Schlumberger, M; Mercken, L; Monier, R; Feunteun, J

    1994-01-01

    Transgenic mice have been used to address the issue of the oncogenic potential of mutant guanine nucleotide stimulatory factor (Gs) alpha subunit in the thyroid gland. The expression of the mutant Arg-201-->His Gs alpha subunit transgene has been directed to murine thyroid epithelial cells by bovine thyroglobulin promoter. The transgenic animals develop hyperfunctioning thyroid adenomas with increased intracellular cAMP levels and high uptake of [125I]iodine and produced elevated levels of circulating triiodothyronine and thyroxine. These animals demonstrate that the mutant form of Gs alpha subunit carries an oncogenic activity, thus supporting the model that deregulation of cAMP level alters growth control in thyroid epithelium. These animals represent models for humans with autonomously functioning thyroid nodules. Images PMID:7937980

  6. Regulation of oncogene-induced cell cycle exit and senescence by chromatin modifiers

    PubMed Central

    David, Gregory

    2012-01-01

    Oncogene activation leads to dramatic changes in numerous biological pathways controlling cellular division, and results in the initiation of a transcriptional program that promotes transformation. Conversely, it also triggers an irreversible cell cycle exit called cellular senescence, which allows the organism to counteract the potentially detrimental uncontrolled proliferation of damaged cells. Therefore, a tight transcriptional control is required at the onset of oncogenic signal, coordinating both positive and negative regulation of gene expression. Not surprisingly, numerous chromatin modifiers contribute to the cellular response to oncogenic stress. While these chromatin modifiers were initially thought of as mere mediators of the cellular response to oncogenic stress, recent studies have uncovered a direct and specific regulation of chromatin modifiers by oncogenic signals. We review here the diverse functions of chromatin modifiers in the cellular response to oncogenic stress, and discuss the implications of these findings on the regulation of cell cycle progression and proliferation by activated oncogenes. PMID:22825329

  7. In vivo photoacoustic imaging of tyrosinase expressing tumours in mice

    NASA Astrophysics Data System (ADS)

    Laufer, Jan; Jathoul, Amit; Johnson, Peter; Zhang, Edward; Lythgoe, Mark; Pedley, R. Barbara; Pule, Martin; Beard, Paul

    2012-02-01

    Two human tumour cell lines (K562, 293T) were stably transfected to achieve the genetic expression of tyrosinase, which is involved in the production of the pigment eumelanin. The cells were injected subcutaneously into nude mice to form tumour xenografts, which were imaged over a period of up to 26 days using an all-optical photoacoustic imaging system. 3D photoacoustic images of the tumours and the surrounding vasculature were acquired at excitation wavelengths ranging from 600nm to 770nm. The images showed tumour growth and continued tyrosinase expression over the full 26 day duration of the study. These findings were confirmed by histological analysis of excised tumour samples.

  8. Genome-Wide Gene Expression Analysis Identifies the Proto-oncogene Tyrosine-Protein Kinase Src as a Crucial Virulence Determinant of Infectious Laryngotracheitis Virus in Chicken Cells

    PubMed Central

    Li, Hai; Wang, Fengjie; Han, Zongxi; Gao, Qi; Li, Huixin; Shao, Yuhao; Sun, Nana

    2015-01-01

    ABSTRACT Given the side effects of vaccination against infectious laryngotracheitis (ILT), novel strategies for ILT control and therapy are urgently needed. The modulation of host-virus interactions is a promising strategy to combat the virus; however, the interactions between the host and avian ILT herpesvirus (ILTV) are unclear. Using genome-wide transcriptome studies in combination with a bioinformatic analysis, we identified proto-oncogene tyrosine-protein kinase Src (Src) to be an important modulator of ILTV infection. Src controls the virulence of ILTV and is phosphorylated upon ILTV infection. Functional studies revealed that Src prolongs the survival of host cells by increasing the threshold of virus-induced cell death. Therefore, Src is essential for viral replication in vitro and in ovo but is not required for ILTV-induced cell death. Furthermore, our results identify a positive-feedback loop between Src and the tyrosine kinase focal adhesion kinase (FAK), which is necessary for the phosphorylation of either Src or FAK and is required for Src to modulate ILTV infection. To the best of our knowledge, we are the first to identify a key host regulator controlling host-ILTV interactions. We believe that our findings have revealed a new potential therapeutic target for ILT control and therapy. IMPORTANCE Despite the extensive administration of live attenuated vaccines starting from the mid-20th century and the administration of recombinant vaccines in recent years, infectious laryngotracheitis (ILT) outbreaks due to avian ILT herpesvirus (ILTV) occur worldwide annually. Presently, there are no drugs or control strategies that effectively treat ILT. Targeting of host-virus interactions is considered to be a promising strategy for controlling ILTV infections. However, little is known about the mechanisms governing host-ILTV interactions. The results from our study advance our understanding of host-ILTV interactions on a molecular level and provide experimental

  9. Platelet-activating factor induces phospholipid turnover, calcium flux, arachidonic acid liberation, eicosanoid generation, and oncogene expression in a human B cell line

    SciTech Connect

    Schulam, P.G.; Kuruvilla, A.; Putcha, G.; Mangus, L.; Franklin-Johnson, J.; Shearer, W.T. )

    1991-03-01

    Platelet-activating factor is a potent mediator of the inflammatory response. Studies of the actions of platelet-activating factor have centered mainly around neutrophils, monocytes, and platelets. In this report we begin to uncover the influence of platelet-activating factor on B lymphocytes. Employing the EBV-transformed human B cell line SKW6.4, we demonstrate that platelet-activating factor significantly alters membrane phospholipid metabolism indicated by the incorporation of 32P into phosphatidylcholine, phosphatidylinositol, and phosphatidic acid but not significantly into phosphatidylethanolamine at concentrations ranging from 10(-9) to 10(-6) M. The inactive precursor, lyso-platelet-activating factor, at a concentration as high as 10(-7) M had no effect on any of the membrane phospholipids. We also show that platelet-activating factor from 10(-12) to 10(-6) M induced rapid and significant elevation in intracellular calcium levels, whereas lyso-platelet-activating factor was again ineffective. We further demonstrate the impact of platelet-activating factor binding to B cells by measuring platelet-activating factor induced arachidonic acid release and 5-hydroxyeicosatetraenoic acid production. Moreover, platelet-activating factor was capable of inducing transcription of the nuclear proto-oncogenes c-fos and c-jun. Finally we explored the possible role of 5-hydroxyeicosatetraenoic acid as a regulator of arachidonic acid liberation demonstrating that endogenous 5-lipoxygenase activity modulates platelet-activating factor induced arachidonic acid release perhaps acting at the level of phospholipase A2. In summary, platelet-activating factor is shown here to have a direct and profound effect on a pure B cell line.

  10. Principles of Cancer Therapy: Oncogene and Non-oncogene Addiction

    PubMed Central

    Luo, Ji; Solimini, Nicole L.; Elledge, Stephen J.

    2010-01-01

    Cancer is a complex collection of distinct genetic diseases united by common hallmarks. Here, we expand upon the classic hallmarks to include the stress phenotypes of tumorigenesis. We describe a conceptual framework of how oncogene and non-oncogene addictions contribute to these hallmarks and how they can be exploited through stress sensitization and stress overload to selectively kill cancer cells. In particular, we present evidence for a large class of non-oncogenes that are essential for cancer cell survival and present attractive drug targets. Finally, we discuss the path ahead to therapeutic discovery and provide theoretical considerations for combining orthogonal cancer therapies. PMID:19269363

  11. Principles of cancer therapy: oncogene and non-oncogene addiction.

    PubMed

    Luo, Ji; Solimini, Nicole L; Elledge, Stephen J

    2009-03-01

    Cancer is a complex collection of distinct genetic diseases united by common hallmarks. Here, we expand upon the classic hallmarks to include the stress phenotypes of tumorigenesis. We describe a conceptual framework of how oncogene and non-oncogene addictions contribute to these hallmarks and how they can be exploited through stress sensitization and stress overload to selectively kill cancer cells. In particular, we present evidence for a large class of non-oncogenes that are essential for cancer cell survival and present attractive drug targets. Finally, we discuss the path ahead to therapeutic discovery and provide theoretical considerations for combining orthogonal cancer therapies. PMID:19269363

  12. Oncogenes in human testicular cancer: DNA and RNA studies.

    PubMed Central

    Peltomäki, P.; Alfthan, O.; de la Chapelle, A.

    1991-01-01

    Oncogene dosage and expression were studied in 16 testicular neoplasms, 14 of germ cell and two of non-germ cell origin. In comparison with normal DNA, tumour DNA of a total of eight patients (seven with germ cell neoplasm and one with testicular lymphoma) showed increased dosages of KRAS2, PDGFA, EGFR, MET and PDGFB. The most frequent (occurring in six tumours) and prominent (up to 3-4-fold) increases were detected in the dosages of KRAS2 (on chromosome 12p) and PDGFA (chromosome 7p), relative to a reference locus from chromosome 2. Importantly, there was a similar increase in 12p dosage in general in these tumours, suggesting the presence of the characteristic isochromosome 12p marker. On the contrary, possible 7p polysomy (assessed by molecular methods) did not explain the PDGFA (or EGFR) changes in all cases. NRAS, MYCN, CSFIR, MYB, MYC, ABL, HRASI, TP53, and ERBB2 did not reveal any consistent alterations in tumour DNA. In RNA dot blot assays the expression of KRAS2, PDGFA, EGFR, or MYC was generally not increased in the tumour samples when compared to that in normal testicular tissue of the same patients although there was interindividual variation in mRNA levels. It thus appears that while oncogene dosage changes occur in a proportion of testis cancers, they are often part of changes in large chromosomal regions or whole arms and are seldom accompanied by altered expression. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:1829952

  13. Pancreatitis promotes oncogenic KrasG12D-induced pancreatic transformation through activation of Nupr1

    PubMed Central

    Grasso, Daniel; Garcia, Maria Noé; Hamidi, Tewfik; Cano, Carla; Calvo, Ezequiel; Lomberk, Gwen; Urrutia, Raul; Iovanna, Juan L

    2014-01-01

    During the initiation stage of pancreatic adenocarcinoma induced by oncogenic Kras, pancreatic cells are exposed to both a protumoral effect and an opposing tumor suppressive process known as oncogene-induced senescence. Pancreatitis disrupts this balance in favor of the transforming effect of oncogenes by lowering the tumor suppressive threshold of oncogene-induced senescence through expression of the stress protein Nupr1. PMID:27308320

  14. The DNA binding site of the Dof protein NtBBF1 is essential for tissue-specific and auxin-regulated expression of the rolB oncogene in plants.

    PubMed Central

    Baumann, K; De Paolis, A; Costantino, P; Gualberti, G

    1999-01-01

    The Dof proteins are a large family of plant transcription factors that share a single highly conserved zinc finger. The tobacco Dof protein NtBBF1 was identified by its ability to bind to regulatory domain B in the promoter of the rolB oncogene. In this study, we show that the ACT T TA target sequence of NtBBF1 in domain B is necessary for tissue-specific expression of rolB. beta-Glucuronidase (GUS) activity of tobacco plants containing a rolB promoter-GUS fusion with a mutated NtBBF1 target sequence within domain B is almost completely suppressed in apical meristems and is severely abated in the vascular system. The ACT T TA motif is shown here also to be one of the cis-regulatory elements involved in auxin induction of rolB. The pattern of NtBBF1 expression in plants is remarkably similar to that of rolB, except in mesophyll cells of mature leaves, in which only NtBBF1 expression could be detected. Ectopic expression of rolB in mesophyll cells was achieved by particle gun delivery if the NtBBF1 binding sequence was intact. These data provide evidence that in the plant, a Dof protein DNA binding sequence acts as a transcriptional regulatory motif, and they point to NtBBF1 as the protein involved in mediating tissue-specific and auxin-inducible expression of rolB. PMID:10072394

  15. Quantitative imaging of gene expression in Drosophila embryos.

    PubMed

    Surkova, Svetlana; Myasnikova, Ekaterina; Kozlov, Konstantin N; Pisarev, Andrei; Reinitz, John; Samsonova, Maria

    2013-06-01

    Quantitative measurements derived using sophisticated microscopy techniques are essential for understanding the basic principles that control the behavior of biological systems. Here we describe a data pipeline developed to extract quantitative data on segmentation gene expression from confocal images of gene expression patterns in Drosophila. The pipeline consists of image segmentation, background removal, temporal characterization of an embryo, data registration, and data averaging. This pipeline has been successfully applied to obtain quantitative gene expression data at cellular resolution in space and at 6.5-min resolution in time. It has also enabled the construction of a spatiotemporal atlas of segmentation gene expression. We describe the software used to construct a workflow for extracting quantitative data on segmentation gene expression and the BREReA package, which implements the methods for background removal and registration of segmentation gene expression patterns. PMID:23734022

  16. The Epstein-Barr virus oncogene product, latent membrane protein 1, induces the downregulation of E-cadherin gene expression via activation of DNA methyltransferases.

    PubMed

    Tsai, Chi-Neu; Tsai, Chia-Lung; Tse, Ka-Po; Chang, Hwan-You; Chang, Yu-Sun

    2002-07-23

    The latent membrane protein (LMP1) of Epstein-Barr virus (EBV) is expressed in EBV-associated nasopharyngeal carcinoma, which is notoriously metastatic. Although it is established that LMP1 represses E-cadherin expression and enhances the invasive ability of carcinoma cells, the mechanism underlying this repression remains to be elucidated. In this study, we demonstrate that LMP1 induces the expression and activity of the DNA methyltransferases 1, 3a, and 3b, using real-time reverse transcription-PCR and enzyme activity assay. This results in hypermethylation of the E-cadherin promoter and down-regulation of E-cadherin gene expression, as revealed by methylation-specific PCR, real-time reverse transcription-PCR and Western blotting data. The DNA methyltransferase inhibitor, 5'-Aza-2'dC, restores E-cadherin promoter activity and protein expression in LMP1-expressing cells, which in turn blocks cell migration ability, as demonstrated by the Transwell cell migration assay. Our findings suggest that LMP1 down-regulates E-cadherin gene expression and induces cell migration activity by using cellular DNA methylation machinery. PMID:12110730

  17. Investigation of Astragalus honey and propolis extract's cytotoxic effect on two human cancer cell lines and their oncogen and proapoptotic gene expression profiles

    PubMed Central

    Sadeghi-Aliabadi, Hojjat; Hamzeh, Jamal; Mirian, Mina

    2015-01-01

    Background: Cancer is one of the major fatal human diseases. Natural products have been used in the treatment of cancer for long time. Bee products including honey and propolis have been introduced for malignancy treatment in recent decades. In this study cytotoxicity of bee products and their effects on the expression of proapoptotic genes have been investigated. Materials and Methods: Cytotoxic effects of Astragalus honey, ethanol extract of propolis and a sugar solution (as control) against HepG2, 5637 and L929 cell lines have been evaluated by the MTT assay. Total RNAs of treated cells were isolated and p53 and Bcl-2 gene expression were evaluated, using real-time PCR. Results: Propolis IC50 values were 58, 30 and 15 μg/ml against L929, HepG2 and 5637, respectively. These values for honey were 3.1%, 2.4% and 1.9%, respectively. Propolis extract has increased the expression of the Bcl-2 gene in all cell lines whereas the honey decreased that significantly (P < 0.05). Also, we found that honey and propolis decreased p53 gene expression in HepG2 and 5637 significantly but not in L929 cells. The sugar solution increased the expression of p53 in two cancer cell lines but no significant changes were observed in the expression of this gene in L929 as normal mouse cell. Conclusion: By downregulation of Bcl-2 expression it could be concluded that the cytotoxicity of honey was more than two fold against tested cancer cells compared with the sugar solution. No significant changes were observed in the expression of p53 in honey-treated cells. Propolis had no significant effect on Bcl-2 and p53 gene expressions (P > 0.05). PMID:25789268

  18. NF-kB and c-Jun induce the expression of the oncogenic miR-221 and miR-222 in prostate carcinoma and glioblastoma cells.

    PubMed

    Galardi, Silvia; Mercatelli, Neri; Farace, Maria G; Ciafrè, Silvia A

    2011-05-01

    MicroRNAs (miRNAs) are potent negative regulators of gene expression involved in all aspects of cell biology. They finely modulate virtually all physiological pathways in metazoans, and are deeply implicated in all main pathologies, among which cancer. Mir-221 and miR-222, two closely related miRNAs encoded in cluster from a genomic region on chromosome X, are strongly upregulated in several forms of human tumours. In this work, we report that the ectopic modulation of NF-kB modifies miR-221/222 expression in prostate carcinoma and glioblastoma cell lines, where we had previously shown their oncogenic activity. We identify two separate distal regions upstream of miR-221/222 promoter which are bound by the NF-kB subunit p65 and drive efficient transcription in luciferase reporter assays; consistently, the site-directed mutagenesis disrupting p65 binding sites or the ectopical inhibition of NF-kB activity significantly reduce luciferase activity. In the most distal enhancer region, we also define a binding site for c-Jun, and we show that the binding of this factor cooperates with that of p65, fully accounting for the observed upregulation of miR-221/222. Thus our work uncovers an additional mechanism through which NF-kB and c-Jun, two transcription factors deeply involved in cancer onset and progression, contribute to oncogenesis, by inducing miR-221/222 transcription. PMID:21245048

  19. NF-kB and c-Jun induce the expression of the oncogenic miR-221 and miR-222 in prostate carcinoma and glioblastoma cells

    PubMed Central

    Galardi, Silvia; Mercatelli, Neri; Farace, Maria G.; Ciafrè, Silvia A.

    2011-01-01

    MicroRNAs (miRNAs) are potent negative regulators of gene expression involved in all aspects of cell biology. They finely modulate virtually all physiological pathways in metazoans, and are deeply implicated in all main pathologies, among which cancer. Mir-221 and miR-222, two closely related miRNAs encoded in cluster from a genomic region on chromosome X, are strongly upregulated in several forms of human tumours. In this work, we report that the ectopic modulation of NF-kB modifies miR-221/222 expression in prostate carcinoma and glioblastoma cell lines, where we had previously shown their oncogenic activity. We identify two separate distal regions upstream of miR-221/222 promoter which are bound by the NF-kB subunit p65 and drive efficient transcription in luciferase reporter assays; consistently, the site-directed mutagenesis disrupting p65 binding sites or the ectopical inhibition of NF-kB activity significantly reduce luciferase activity. In the most distal enhancer region, we also define a binding site for c-Jun, and we show that the binding of this factor cooperates with that of p65, fully accounting for the observed upregulation of miR-221/222. Thus our work uncovers an additional mechanism through which NF-kB and c-Jun, two transcription factors deeply involved in cancer onset and progression, contribute to oncogenesis, by inducing miR-221/222 transcription. PMID:21245048

  20. Leucine leucine-37 uses formyl peptide receptor-like 1 to activate signal transduction pathways, stimulate oncogenic gene expression, and enhance the invasiveness of ovarian cancer cells.

    PubMed

    Coffelt, Seth B; Tomchuck, Suzanne L; Zwezdaryk, Kevin J; Danka, Elizabeth S; Scandurro, Aline B

    2009-06-01

    Emerging evidence suggests that the antimicrobial peptide, leucine leucine-37 (LL-37), could play a role in the progression of solid tumors. LL-37 is expressed as the COOH terminus of human cationic antimicrobial protein-18 (hCAP-18) in ovarian, breast, and lung cancers. Previous studies have shown that the addition of LL-37 to various cancer cell lines in vitro stimulates proliferation, migration, and invasion. Similarly, overexpression of hCAP-18/LL-37 in vivo accelerates tumor growth. However, the receptor or receptors through which these processes are mediated have not been thoroughly examined. In the present study, expression of formyl peptide receptor-like 1 (FPRL1) was confirmed on ovarian cancer cells. Proliferation assays indicated that LL-37 does not signal through a G protein-coupled receptor, such as FPRL1, to promote cancer cell growth. By contrast, FPRL1 was required for LL-37-induced invasion through Matrigel. The peptide stimulated mitogen-activated protein kinase and Janus-activated kinase/signal transducers and activators of transcription signaling cascades and led to the significant activation of several transcription factors, through both FPRL1-dependent and FPRL1-independent pathways. Likewise, expression of some LL-37-stimulated genes was attenuated by the inhibition of FPRL1. Increased expression of CXCL10, EGF, and PDGF-BB as well as other soluble factors was confirmed from conditioned medium of LL-37-treated cells. Taken together, these data suggest that LL-37 potentiates a more aggressive behavior from ovarian cancer cells through its interaction with FPRL1. PMID:19491199

  1. Leucine Leucine-37 Uses Formyl Peptide Receptor–Like 1 to Activate Signal Transduction Pathways, Stimulate Oncogenic Gene Expression, and Enhance the Invasiveness of Ovarian Cancer Cells

    PubMed Central

    Coffelt, Seth B.; Tomchuck, Suzanne L.; Zwezdaryk, Kevin J.; Danka, Elizabeth S.; Scandurro, Aline B.

    2009-01-01

    Emerging evidence suggests that the antimicrobial peptide, leucine leucine-37 (LL-37), could play a role in the progression of solid tumors. LL-37 is expressed as the COOH terminus of human cationic antimicrobial protein-18 (hCAP-18) in ovarian, breast, and lung cancers. Previous studies have shown that the addition of LL-37 to various cancer cell lines in vitro stimulates proliferation, migration, and invasion. Similarly, overexpression of hCAP-18/LL-37 in vivo accelerates tumor growth. However, the receptor or receptors through which these processes are mediated have not been thoroughly examined. In the present study, expression of formyl peptide receptor–like 1 (FPRL1) was confirmed on ovarian cancer cells. Proliferation assays indicated that LL-37 does not signal through a G protein–coupled receptor, such as FPRL1, to promote cancer cell growth. By contrast, FPRL1 was required for LL-37–induced invasion through Matrigel. The peptide stimulated mitogen-activated protein kinase and Janus-activated kinase/signal transducers and activators of transcription signaling cascades and led to the significant activation of several transcription factors, through both FPRL1-dependent and FPRL1-independent pathways. Likewise, expression of some LL-37–stimulated genes was attenuated by the inhibition of FPRL1. Increased expression of CXCL10, EGF, and PDGF-BB as well as other soluble factors was confirmed from conditioned medium of LL-37–treated cells. Taken together, these data suggest that LL-37 potentiates a more aggressive behavior from ovarian cancer cells through its interaction with FPRL1. PMID:19491199

  2. Oncogenic role of nucleophosmin/B23.

    PubMed

    Yung, Benjamin Yat Ming

    2007-01-01

    Nucleophosmin/B23 was first identified as a nucleolar protein expressed at higher levels in cancer cells compared to normal cells. Nucleophosmin/B23 has long been thus thought to have a role in tumor formation. With our efforts and others in the last 15 years, nucleophosmin/B23 has proven to have an oncogenic role. In this review, we provide evidence suggesting that nucleophosmin/B23 may be a crucial gene in regulation of cancer growth and discuss how nucleophosmin/B23 can contribute to tumorigenesis. PMID:17939258

  3. Discrimination of gender using facial image with expression change

    NASA Astrophysics Data System (ADS)

    Kuniyada, Jun; Fukuda, Takahiro; Terada, Kenji

    2005-12-01

    By carrying out marketing research, the managers of large-sized department stores or small convenience stores obtain the information such as ratio of men and women of visitors and an age group, and improve their management plan. However, these works are carried out in the manual operations, and it becomes a big burden to small stores. In this paper, the authors propose a method of men and women discrimination by extracting difference of the facial expression change from color facial images. Now, there are a lot of methods of the automatic recognition of the individual using a motion facial image or a still facial image in the field of image processing. However, it is very difficult to discriminate gender under the influence of the hairstyle and clothes, etc. Therefore, we propose the method which is not affected by personality such as size and position of facial parts by paying attention to a change of an expression. In this method, it is necessary to obtain two facial images with an expression and an expressionless. First, a region of facial surface and the regions of facial parts such as eyes, nose, and mouth are extracted in the facial image with color information of hue and saturation in HSV color system and emphasized edge information. Next, the features are extracted by calculating the rate of the change of each facial part generated by an expression change. In the last step, the values of those features are compared between the input data and the database, and the gender is discriminated. In this paper, it experimented for the laughing expression and smile expression, and good results were provided for discriminating gender.

  4. Imaging Axl expression in pancreatic and prostate cancer xenografts

    SciTech Connect

    Nimmagadda, Sridhar; Pullambhatla, Mrudula; Lisok, Ala; Hu, Chaoxin; Maitra, Anirban; Pomper, Martin G

    2014-01-10

    Highlights: •Axl is overexpressed in a variety of cancers. •Axl overexpression confers invasive phenotype. •Axl imaging would be useful for therapeutic guidance and monitoring. •Axl expression imaging is demonstrated in pancreatic and prostate cancer xenografts. •Graded levels of Axl expression imaging is feasible. -- Abstract: The receptor tyrosine kinase Axl is overexpressed in and leads to patient morbidity and mortality in a variety of cancers. Axl–Gas6 interactions are critical for tumor growth, angiogenesis and metastasis. The goal of this study was to investigate the feasibility of imaging graded levels of Axl expression in tumors using a radiolabeled antibody. We radiolabeled anti-human Axl (Axl mAb) and control IgG1 antibodies with {sup 125}I with high specific radioactivity and radiochemical purity, resulting in an immunoreactive fraction suitable for in vivo studies. Radiolabeled antibodies were investigated in severe combined immunodeficient mice harboring subcutaneous CFPAC (Axl{sup high}) and Panc1 (Axl{sup low}) pancreatic cancer xenografts by ex vivo biodistribution and imaging. Based on these results, the specificity of [{sup 125}I]Axl mAb was also validated in mice harboring orthotopic Panc1 or CFPAC tumors and in mice harboring subcutaneous 22Rv1 (Axl{sup low}) or DU145 (Axl{sup high}) prostate tumors by ex vivo biodistribution and imaging studies at 72 h post-injection of the antibody. Both imaging and biodistribution studies demonstrated specific and persistent accumulation of [{sup 125}I]Axl mAb in Axl{sup high} (CFPAC and DU145) expression tumors compared to the Axl{sup low} (Panc1 and 22Rv1) expression tumors. Axl expression in these tumors was further confirmed by immunohistochemical studies. No difference in the uptake of radioactivity was observed between the control [{sup 125}I]IgG1 antibody in the Axl{sup high} and Axl{sup low} expression tumors. These data demonstrate the feasibility of imaging Axl expression in pancreatic

  5. GENES FOR TUMOR MARKERS ARE CLUSTERED WITH CELLULAR PROTO-ONCOGENES ON HUMAN CHROMOSOMES

    EPA Science Inventory

    The relative mapping positions of genes for polypeptides expressed abnormally in tumors (tumor markers) and cellular proto-oncogenes were analyzed and a remarkable degree of co-mapping of tumor marker genes with oncogenes in the human karyotype were found. It is proposed that abe...

  6. Oncogenes and RNA splicing of human tumor viruses.

    PubMed

    Ajiro, Masahiko; Zheng, Zhi-Ming

    2014-09-01

    Approximately 10.8% of human cancers are associated with infection by an oncogenic virus. These viruses include human papillomavirus (HPV), Epstein-Barr virus (EBV), Merkel cell polyomavirus (MCV), human T-cell leukemia virus 1 (HTLV-1), Kaposi's sarcoma-associated herpesvirus (KSHV), hepatitis C virus (HCV) and hepatitis B virus (HBV). These oncogenic viruses, with the exception of HCV, require the host RNA splicing machinery in order to exercise their oncogenic activities, a strategy that allows the viruses to efficiently export and stabilize viral RNA and to produce spliced RNA isoforms from a bicistronic or polycistronic RNA transcript for efficient protein translation. Infection with a tumor virus affects the expression of host genes, including host RNA splicing factors, which play a key role in regulating viral RNA splicing of oncogene transcripts. A current prospective focus is to explore how alternative RNA splicing and the expression of viral oncogenes take place in a cell- or tissue-specific manner in virus-induced human carcinogenesis. PMID:26038756

  7. c-fos oncogene underexpression in salivary gland tumors as measured by in situ hybridization.

    PubMed Central

    Birek, C.; Lui, E.; Dardick, I.

    1993-01-01

    Tissue from 35 salivary gland tumors and 14 normal salivary glands was analyzed by in situ hybridization and computer-assisted morphometry for the expression of the c-fos oncogene. The normal salivary gland tissues were found to express c-fos focally, mainly in the acinar secretory cells. The majority of the cells in the normal tissues showed a high level of expression (47.74 +/- 5.31% of cells had 46 to 60 grains per cell and another 45.79 +/- 2.18% showed > 60 grains per cell). All the tumors examined exhibited a relatively low, uniform distribution of c-fos expression. For example, in the poorly differentiated adenocarcinomas, 96.83 +/- 04% of the cells were found to have < 15 grains per cell. A general linear model for multivariate analysis showed a significant difference between the various tumor types and the normal salivary gland tissues (P = 0.0001). These data support the hypothesis that salivary gland tumors belong to a group of epithelial neoplasias in which the loss of cellular differentiation is linked with underexpression of the c-fos oncogene. Images Figure 1 Figure 2 Figure 3 PMID:8456948

  8. Differential protein expression and oncogenic gene network link tyrosine kinase ephrin B4 receptor to aggressive gastric and gastroesophageal junction cancers.

    PubMed

    Liersch-Löhn, Britta; Slavova, Nadia; Buhr, Heinz J; Bennani-Baiti, Idriss M

    2016-03-01

    Transmembrane tyrosine-kinase Ephrin receptors promote tumor progression and/or metastasis of several malignancies including leukemia, follicular lymphoma, glioma, malignant pleural mesothelioma, papillary thyroid carcinoma, sarcomas and ovarian, breast, bladder and non-small cell lung cancers. They also drive intestinal stem cell proliferation and positioning, control intestinal tissue boundaries and are involved in liver, pancreatic and colorectal cancers, indicating involvement in additional digestive system malignancies. We investigated the role of Ephrin-B4 receptor (EPHB4), and its ligand EFNB2, in gastric and gastroesophageal junction cancers in patient cohorts through computational, mathematical, molecular and immunohistochemical analyses. We show that EPHB4 is upregulated in preneoplastic gastroesophageal lesions and its expression further increased in gastroesophageal cancers in several independent cohorts. The closely related EPHB6 receptor, which also binds EFNB2, was downregulated in all tested cohorts, consistent with its tumor-suppressive properties in other cancers. EFNB2 expression is induced in esophageal cells by acidity, suggesting that gastroesophageal reflux disease (GERD) may constitute an early triggering event in activating EFNB2-EPHB4 signaling. Association of EPHB4 to both Barrett's esophagus and to advanced tumor stages, and its overexpression at the tumor invasion front and vascular endothelial cells intimate the notion that EPHB4 may be associated with multiple steps of gastroesophageal tumorigenesis. Analysis of oncogenomic signatures uncovered the first EPHB4-associated gene network (false discovery rate: 7 × 10(-90) ) composed of a five-transcription factor interconnected gene network that drives proliferation, angiogenesis and invasiveness. The EPHB4 oncogenomic network provides a molecular basis for its role in tumor progression and points to EPHB4 as a potential tumor aggressiveness biomarker and drug target in gastroesophageal

  9. Imaging of gene expression in vivo with photoacoustic tomography

    NASA Astrophysics Data System (ADS)

    Li, Li; Zemp, Roger J.; Lungu, Gina; Stoica, George; Wang, Lihong V.

    2006-02-01

    In the post-genomic era, there is an increasing interest in visualizing the expression of functional genes in vivo. With the assistance of the reporter gene technique, various imaging modalities have been adopted for this purpose. In vivo gene expression imaging promises to provide biologists with a powerful tool for deepening our understanding of developmental biology, expanding our knowledge of the genetic basis of disease, and advancing the development of medicine. In this paper, we demonstrate the feasibility of imaging gene expression with photoacoustic imaging, which offers unique absorption contrast with ultrasonic resolution in vivo. We mark tumors in rats with the lacZ reporter gene. The lacZ gene encodes an enzyme β-galactosidase, which yields a dark blue product when acting on a colorimetric assay called X-gal. Photoacoustic tomography at 650nm clearly visualizes the presence of this blue product. The spectroscopic method can also potentially improve specificity. Considering how many staining methods are used in traditional biology, we believe that photoacoustic techniques will revolutionize the field of molecular imaging. The further development of reporter gene systems with high absorbing products in the NIR region is needed.

  10. TPR-MET oncogenic rearrangement: Detection by polymerase chain reaction amplification of the transcript and expression in human tumor cells lines

    SciTech Connect

    Soman, N.R.; Wogan, G.N. ); Rhim, J.S. )

    1990-01-01

    Activation of the MET protooncogene by a rearrangement involving the fusion of TPR and MET specific gene sequences has been observed in a human osteosarcoma cell line (HOS) treated in vitro with N-methyl-N{prime}-nitro-N-nitrosoguanidine (MNNG). No information has been available about the possible occurrence of this rearrangement in human tumors. To facilitate rapid screening of human cell lines and tumor samples for this specific gene rearrangement; the authors developed a sensitive detection method based on polymerase chain reaction (PCR) amplification of TPR-MET mRNA. cDNA was generated from cellular transcripts by using one of the PCR primers, which was then used as a template for PCR amplification of a 205-base-pair region carrying the breakpoint. An end-labeled internal probe was hybridized in solution to an aliquot of the PCR product for detecting amplification. Cells could be directly screened by the assay without prior isolation of RNA. A 205-base-pair DNA fragment characteristic of the TRP-MET rearrangement was detected in cell lines previously known to contain this altered sequence. The rearrangement was also detected at very low levels in the parental (nontransformed) cell line, HOS TE-85. A preliminary survey of cell lines derived from a variety of human tumors indicates that TPR-MET rearrangement occurred and was expressed at very low frequencies by cells from 7 of 14 tumors of nonhematopoietic origin.

  11. Interferon-induced revertants of ras-transformed cells: resistance to transformation by specific oncogenes and retransformation by 5-azacytidine.

    PubMed Central

    Samid, D; Flessate, D M; Friedman, R M

    1987-01-01

    Prolonged alpha/beta interferon (IFN-alpha/beta) treatment of NIH 3T3 cells transformed by a long terminal repeat-activated Ha-ras proto-oncogene resulted in revertants that maintained a nontransformed phenotype long after IFN treatment had been discontinued. Cloned persistent revertants (PRs) produced large amounts of the ras-encoded p21 and were refractile to transformation by EJras DNA and by transforming retroviruses which carried the v-Ha-ras, v-Ki-ras, v-abl, or v-fes oncogene. Transient treatment either in vitro or in vivo with cytidine analogs that alter gene expression by inhibiting DNA methylation resulted in transformation of PR, but not of NIH 3T3, cells. The PR retransformants reverted again with IFN, suggesting that DNA methylation is involved in IFN-induced persistent reversion. Images PMID:2439904

  12. miR-17–92 explains MYC oncogene addiction

    PubMed Central

    Li, Yulin; Casey, Stephanie C; Choi, Peter S; Felsher, Dean W

    2014-01-01

    MYC regulates tumorigenesis by coordinating the expression of thousands of genes. We found that MYC appears to regulate the decisions between cell survival versus death and self-renewal versus senescence through the microRNA miR-17–92 cluster. Addiction to the MYC oncogene may therefore in fact be an addiction to miR-17–92. PMID:27308380

  13. Photodynamic treatment (ALA-PDT) suppresses the expression of the oncogenic Bcr-Abl kinase and affects the cytoskeleton organization in K562 cells.

    PubMed

    Pluskalová, Michaela; Peslová, Gabriela; Grebenová, Dana; Halada, Petr; Hrkal, Zbynek

    2006-06-01

    K562 is the chronic myelogenous leukemia (CML)-derived cell line that expresses high levels of chimeric oncoprotein Bcr-Abl. The deregulated (permanent) kinase activity of Bcr-Abl leads to continuous proliferation of K562 cells and their resistance to the apoptosis promotion by conventional drugs. The photodynamic treatment (PDT) based on the application of 5-aminolevulinic acid (ALA) and irradiation with blue light (ALA-PDT) resulted in the suppression of K562 cells proliferation. It was followed by a necrosis-like cell death [K. Kuzelová, D. Grebenová, M. Pluskalová, I. Marinov, Z. Hrkal, J. Photochem. Photobiol. B 73 (2004) 67-78]. ALA-PDT led to the perturbation of the Hsp90/p23 multichaperone complex of which the Bcr-Abl is the client protein. Bcr-Abl protein was suppressed whereas the bcr-abl mRNA level was not affected. Further on, we observed several changes in the cytoskeleton organization. We detected ALA-PDT-mediated disruption of filamental actin structure using FITC-Phalloidin staining. In connection with this we uncovered certain cytoskeleton organizing proteins involved in the cell response to the treatment. Among these proteins, Septin2, which plays a role in maintaining actin bundles, was suppressed. Another one, PDZ-LIM domain protein 1 (CLP36) was altered. This protein acts as an adaptor molecule for LIM-kinase which phosphorylates and thus inactivates cofilin. Cofilin was indeed dephosphorylated and could thus be activated and operate as an actin-depolymerizing factor. We propose the scheme of molecular response of K562 cells to ALA-PDT. PMID:16495075

  14. Oncogenic Ras/Src cooperativity in pancreatic neoplasia

    PubMed Central

    Shields, DJ; Murphy, EA; Desgrosellier, JS; Mielgo, A; Lau, SKM; Barnes, LA; Lesperance, J; Huang, M; Schmedt, C; Tarin, D; Lowy, AM; Cheresh, DA

    2011-01-01

    Pancreas cancer is one of the most lethal malignancies and is characterized by activating mutations of Kras, present in 95% of patients. More than 60% of pancreatic cancers also display increased c-Src activity, which is associated with poor prognosis. Although loss of tumor suppressor function (for example, p16, p53, Smad4) combined with oncogenic Kras signaling has been shown to accelerate pancreatic duct carcinogenesis, it is unclear whether elevated Src activity contributes to Kras-dependent tumorigenesis or is simply a biomarker of disease progression. Here, we demonstrate that in the context of oncogenic Kras, activation of c-Src through deletion of C-terminal Src kinase (CSK) results in the development of invasive pancreatic ductal adenocarcinoma (PDA) by 5–8 weeks. In contrast, deletion of CSK alone fails to induce neoplasia, while oncogenic Kras expression yields PDA at low frequency after a latency of 12 months. Analysis of cell lines derived from Ras/Src-induced PDA’s indicates that oncogenic Ras/Src cooperativity may lead to genomic instability, yet Ras/Src-driven tumor cells remain dependent on Src signaling and as such, Src inhibition suppresses growth of Ras/Src-driven tumors. These findings demonstrate that oncogenic Ras/Src cooperate to accelerate PDA onset and support further studies of Src-directed therapies in pancreatic cancer. PMID:21242978

  15. Oncogene regulation. An oncogenic super-enhancer formed through somatic mutation of a noncoding intergenic element.

    PubMed

    Mansour, Marc R; Abraham, Brian J; Anders, Lars; Berezovskaya, Alla; Gutierrez, Alejandro; Durbin, Adam D; Etchin, Julia; Lawton, Lee; Sallan, Stephen E; Silverman, Lewis B; Loh, Mignon L; Hunger, Stephen P; Sanda, Takaomi; Young, Richard A; Look, A Thomas

    2014-12-12

    In certain human cancers, the expression of critical oncogenes is driven from large regulatory elements, called super-enhancers, that recruit much of the cell's transcriptional apparatus and are defined by extensive acetylation of histone H3 lysine 27 (H3K27ac). In a subset of T-cell acute lymphoblastic leukemia (T-ALL) cases, we found that heterozygous somatic mutations are acquired that introduce binding motifs for the MYB transcription factor in a precise noncoding site, which creates a super-enhancer upstream of the TAL1 oncogene. MYB binds to this new site and recruits its H3K27 acetylase-binding partner CBP, as well as core components of a major leukemogenic transcriptional complex that contains RUNX1, GATA-3, and TAL1 itself. Additionally, most endogenous super-enhancers found in T-ALL cells are occupied by MYB and CBP, which suggests a general role for MYB in super-enhancer initiation. Thus, this study identifies a genetic mechanism responsible for the generation of oncogenic super-enhancers in malignant cells. PMID:25394790

  16. Mouse Elk oncogene maps to chromosome X and a novel Elk oncogene (Elk3) maps to chromosome 10.

    PubMed

    Tamai, Y; Taketo, M; Nozaki, M; Seldin, M F

    1995-03-20

    The Elk protein is a member of the Ets family found in both vertebrates and invertebrates. Human ELK1 encoded by ELK1 binds alone or together with serum response factor to DNA and regulates gene expression in a variety of biological processes. Using a panel of interspecific backcross mice, we have mapped the Elk oncogene (Elk) and a novel type Elk oncogene (Elk3), closely related to ELK1. Elk maps to Chr X, and Elk3 maps to the proximal region of Chr 10. PMID:7601474

  17. Mouse Elk oncogene maps to chromosome X and a novel Elk oncogene (Elk3) maps to chromosome 10

    SciTech Connect

    Tamai, Yoshitaka; Taketo, Makoto; Nozaki, Masami

    1995-03-20

    The Elk protein is a member of the Ets family found in both vertebrates and invertebrates. Human ELK1 encoded by ELK1 binds alone or together with serum response factor to DNA and regulates gene expression in a variety of biological processes. Using a panel of interspecific backcross mice, we have mapped the Elk oncogene (Elk) and a novel type Elk oncogene (Elk3), closely related to ELK1. Elk maps to Chr X, and Elk3 maps to the proximal region of Chr 10. 18 refs., 1 fig., 1 tab.

  18. TGIF function in oncogenic Wnt signaling.

    PubMed

    Razzaque, Mohammed S; Atfi, Azeddine

    2016-04-01

    Transforming growth-interacting factor (TGIF) has been implicated in the pathogenesis of many types of human cancer, but the underlying mechanisms remained mostly enigmatic. Our recent study has revealed that TGIF functions as a mediator of oncogenic Wnt/β-catenin signaling. We found that TGIF can interact with and sequesters Axin1 and Axin2 into the nucleus, thereby culminating in disassembly of the β-catenin-destruction complex and attendant accumulation of β-catenin in the nucleus, where it activates expression of Wnt target genes, including TGIF itself. We have provided proof-of-concept evidences that high levels of TGIF expression correlate with poor prognosis in patients with triple negative breast cancer (TNBC), and that TGIF empowers Wnt-driven mammary tumorigenesis in vivo. Here, we will briefly summarize how TGIF influences Wnt signaling to promote tumorigenesis. PMID:26522669

  19. Image analysis tools and emerging algorithms for expression proteomics

    PubMed Central

    English, Jane A.; Lisacek, Frederique; Morris, Jeffrey S.; Yang, Guang-Zhong; Dunn, Michael J.

    2012-01-01

    Since their origins in academic endeavours in the 1970s, computational analysis tools have matured into a number of established commercial packages that underpin research in expression proteomics. In this paper we describe the image analysis pipeline for the established 2-D Gel Electrophoresis (2-DE) technique of protein separation, and by first covering signal analysis for Mass Spectrometry (MS), we also explain the current image analysis workflow for the emerging high-throughput ‘shotgun’ proteomics platform of Liquid Chromatography coupled to MS (LC/MS). The bioinformatics challenges for both methods are illustrated and compared, whilst existing commercial and academic packages and their workflows are described from both a user’s and a technical perspective. Attention is given to the importance of sound statistical treatment of the resultant quantifications in the search for differential expression. Despite wide availability of proteomics software, a number of challenges have yet to be overcome regarding algorithm accuracy, objectivity and automation, generally due to deterministic spot-centric approaches that discard information early in the pipeline, propagating errors. We review recent advances in signal and image analysis algorithms in 2-DE, MS, LC/MS and Imaging MS. Particular attention is given to wavelet techniques, automated image-based alignment and differential analysis in 2-DE, Bayesian peak mixture models and functional mixed modelling in MS, and group-wise consensus alignment methods for LC/MS. PMID:21046614

  20. Regulation of protein kinase C activity in neuronal differentiation induced by the N-ras oncogene in PC-12 cells.

    PubMed Central

    Lacal, J C; Cuadrado, A; Jones, J E; Trotta, R; Burstein, D E; Thomson, T; Pellicer, A

    1990-01-01

    Expression of the N-ras oncogene under the control of the glucocorticoid-responsive promoter in the pheochromocytoma cell line UR61, a subline of PC-12 cells, has been used to investigate the differentiation process to neuronal cells triggered by ras oncogenes (I. Guerrero, A. Pellicer, and D. E. Burstein, Biochem. Biophys. Res. Commun. 150:1185-1192, 1988). Using ras-inducible cell lines, we observed that expression of the oncogenic N-ras p21 protein interferes with the ability of phorbol esters to induce downregulation of protein kinase C. This effect was associated with the appearance of immunologically detectable protein kinase C as well as the activity of the enzyme as analyzed either by binding of [3H]phorbol-12,13-dibutyrate in intact cells or by in vitro kinase activity. These results indicate a relationship between ras p21 and protein kinase C in neuronal differentiation in this model system. Comparison to the murine fibroblast system suggests that this relationship may be functional. Images PMID:2188105

  1. Targeting oncogenes to improve breast cancer chemotherapy.

    PubMed

    Christensen, Laura A; Finch, Rick A; Booker, Adam J; Vasquez, Karen M

    2006-04-15

    Despite recent advances in treatment, breast cancer remains a serious health threat for women. Traditional chemotherapies are limited by a lack of specificity for tumor cells and the cell cycle dependence of many chemotherapeutic agents. Here we report a novel strategy to help overcome these limitations. Using triplex-forming oligonucleotides (TFOs) to direct DNA damage site-specifically to oncogenes overexpressed in human breast cancer cells, we show that the effectiveness of the anticancer nucleoside analogue gemcitabine can be improved significantly. TFOs targeted to the promoter region of c-myc directly inhibited gene expression by approximately 40%. When used in combination, specific TFOs increased the incorporation of gemcitabine at the targeted site approximately 4-fold, presumably due to induction of replication-independent DNA synthesis. Cells treated with TFOs and gemcitabine in combination showed a reduction in both cell survival and capacity for anchorage-independent growth (approximately 19% of untreated cells). This combination affected the tumorigenic potential of these cancer cells to a significantly greater extent than either treatment alone. This novel strategy may be used to increase the range of effectiveness of antitumor nucleosides in any tumor which overexpresses a targetable oncogene. Multifaceted chemotherapeutic approaches such as this, coupled with triplex-directed gene targeting, may lead to more than incremental improvements in nonsurgical treatment of breast tumors. PMID:16618728

  2. Imaging gene expression in real-time using aptamers

    SciTech Connect

    Shin, Il Chung

    2011-01-01

    Signal transduction pathways are usually activated by external stimuli and are transient. The downstream changes such as transcription of the activated genes are also transient. Real-time detection of promoter activity is useful for understanding changes in gene expression, especially during cell differentiation and in development. A simple and reliable method for viewing gene expression in real time is not yet available. Reporter proteins such as fluorescent proteins and luciferase allow for non-invasive detection of the products of gene expression in living cells. However, current reporter systems do not provide for real-time imaging of promoter activity in living cells. This is because of the long time period after transcription required for fluorescent protein synthesis and maturation. We have developed an RNA reporter system for imaging in real-time to detect changes in promoter activity as they occur. The RNA reporter uses strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags), which can be expressed from a promoter of choice. The tobramycin, neomycin and PDC RNA aptamers have been utilized for this system and expressed in yeast from the GAL1 promoter. The IMAGEtag RNA kinetics were quantified by RT-qPCR. In yeast precultured in raffinose containing media the GAL1 promoter responded faster than in yeast precultured in glucose containing media. IMAGEtag RNA has relatively short half-life (5.5 min) in yeast. For imaging, the yeast cells are incubated with their ligands that are labeled with fluorescent dyes. To increase signal to noise, ligands have been separately conjugated with the FRET (Förster resonance energy transfer) pairs, Cy3 and Cy5. With these constructs, the transcribed aptamers can be imaged after activation of the promoter by galactose. FRET was confirmed with three different approaches, which were sensitized emission, acceptor photobleaching and donor lifetime by FLIM (fluorescence lifetime imaging

  3. Imaging gene expression in real-time using aptamers

    SciTech Connect

    Shin, Ilchung

    2012-01-01

    Signal transduction pathways are usually activated by external stimuli and are transient. The downstream changes such as transcription of the activated genes are also transient. Real-time detection of promoter activity is useful for understanding changes in gene expression, especially during cell differentiation and in development. A simple and reliable method for viewing gene expression in real time is not yet available. Reporter proteins such as fluorescent proteins and luciferase allow for non-invasive detection of the products of gene expression in living cells. However, current reporter systems do not provide for real-time imaging of promoter activity in living cells. This is because of the long time period after transcription required for fluorescent protein synthesis and maturation. We have developed an RNA reporter system for imaging in real-time to detect changes in promoter activity as they occur. The RNA reporter uses strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags), which can be expressed from a promoter of choice. The tobramycin, neomycin and PDC RNA aptamers have been utilized for this system and expressed in yeast from the GAL1 promoter. The IMAGEtag RNA kinetics were quantified by RT-qPCR. In yeast precultured in raffinose containing media the GAL1 promoter responded faster than in yeast precultured in glucose containing media. IMAGEtag RNA has relatively short half-life (5.5 min) in yeast. For imaging, the yeast cells are incubated with their ligands that are labeled with fluorescent dyes. To increase signal to noise, ligands have been separately conjugated with the FRET (Förster resonance energy transfer) pairs, Cy3 and Cy5. With these constructs, the transcribed aptamers can be imaged after activation of the promoter by galactose. FRET was confirmed with three different approaches, which were sensitized emission, acceptor photobleaching and donor lifetime by FLIM (fluorescence lifetime imaging

  4. Autophagic activity dictates the cellular response to oncogenic RAS

    PubMed Central

    Wang, Yihua; Wang, Xiao Dan; Lapi, Eleonora; Sullivan, Alexandra; Jia, Wei; He, You-Wen; Ratnayaka, Indrika; Zhong, Shan; Goldin, Robert D.; Goemans, Christoph G.; Tolkovsky, Aviva M.; Lu, Xin

    2012-01-01

    RAS is frequently mutated in human cancers and has opposing effects on autophagy and tumorigenesis. Identifying determinants of the cellular responses to RAS is therefore vital in cancer research. Here, we show that autophagic activity dictates the cellular response to oncogenic RAS. N-terminal Apoptosis-stimulating of p53 protein 2 (ASPP2) mediates RAS-induced senescence and inhibits autophagy. Oncogenic RAS-expressing ASPP2(Δ3/Δ3) mouse embryonic fibroblasts that escape senescence express a high level of ATG5/ATG12. Consistent with the notion that autophagy levels control the cellular response to oncogenic RAS, overexpressing ATG5, but not autophagy-deficient ATG5 mutant K130R, bypasses RAS-induced senescence, whereas ATG5 or ATG3 deficiency predisposes to it. Mechanistically, ASPP2 inhibits RAS-induced autophagy by competing with ATG16 to bind ATG5/ATG12 and preventing ATG16/ATG5/ATG12 formation. Hence, ASPP2 modulates oncogenic RAS-induced autophagic activity to dictate the cellular response to RAS: to proliferate or senesce. PMID:22847423

  5. Folate levels modulate oncogene-induced replication stress and tumorigenicity

    PubMed Central

    Lamm, Noa; Maoz, Karin; Bester, Assaf C; Im, Michael M; Shewach, Donna S; Karni, Rotem; Kerem, Batsheva

    2015-01-01

    Chromosomal instability in early cancer stages is caused by replication stress. One mechanism by which oncogene expression induces replication stress is to drive cell proliferation with insufficient nucleotide levels. Cancer development is driven by alterations in both genetic and environmental factors. Here, we investigated whether replication stress can be modulated by both genetic and non-genetic factors and whether the extent of replication stress affects the probability of neoplastic transformation. To do so, we studied the effect of folate, a micronutrient that is essential for nucleotide biosynthesis, on oncogene-induced tumorigenicity. We show that folate deficiency by itself leads to replication stress in a concentration-dependent manner. Folate deficiency significantly enhances oncogene-induced replication stress, leading to increased DNA damage and tumorigenicity in vitro. Importantly, oncogene-expressing cells, when grown under folate deficiency, exhibit a significantly increased frequency of tumor development in mice. These findings suggest that replication stress is a quantitative trait affected by both genetic and non-genetic factors and that the extent of replication stress plays an important role in cancer development. PMID:26197802

  6. Dimerization mediated through a leucine zipper activates the oncogenic potential of the met receptor tyrosine kinase.

    PubMed Central

    Rodrigues, G A; Park, M

    1993-01-01

    Oncogenic activation of the met (hepatocyte growth factor/scatter factor) receptor tyrosine kinase involves a genomic rearrangement that generates a hybrid protein containing tpr-encoded sequences at its amino terminus fused directly to the met-encoded receptor kinase domain. Deletion of Tpr sequences abolishes the transforming ability of this protein, implicating this region in oncogenic activation. We demonstrate, by site-directed mutagenesis and coimmunoprecipitation experiments, that a leucine zipper motif within Tpr mediates dimerization of the tpr-met product and is essential for the transforming activity of the met oncogene. By analogy with ligand-stimulated activation of receptor tyrosine kinases, we propose that constitutive dimerization mediated by a leucine zipper motif within Tpr is responsible for oncogenic activation of the Met kinase. The possibility that this mechanism of activation represents a paradigm for a class of receptor tyrosine kinase oncogenes activated by DNA rearrangement is discussed. Images PMID:8413267

  7. Oncogenicity of human N-ras oncogene and proto-oncogene introduced into retroviral vectors

    SciTech Connect

    Souyri, M.; Vigon, I.; Charon, M.; Tambourin, P. )

    1989-09-01

    The N-ras gene is the only member of the ras family which has never been naturally transduced into a retrovirus. In order to study the in vitro and in vivo oncogenicity of N-ras and to compare its pathogenicity to that of H-ras, the authors have inserted an activated or a normal form of human N-ras cDNA into a slightly modified Harvey murine sarcoma virus-derived vector in which the H-ras p21 coding region had been deleted. The resulting constructions were transfected into NIH 3T3 cells. The activated N-ras-containing construct (HSN) induced 10{sup 4} foci per {mu}g of DNA and was found to be as transforming as H-ras was. After infection of the transfected cells by either the ecotropic Moloney murine leukemia virus or the amphotropic 4070A helper viruses, rescued transforming viruses were injected into newborn mice. Both pseudotypes of HSN virus containing activated N-ras induced the typical Harvey disease with similar latency. However, they found that the virus which contained normal N-ras p21 (HSn) was also pathogenic and induced splenomegaly, lymphadenopathies, and sarcoma in mice after a latency of 3 to 7 weeks. In addition, Moloney murine leukemia virus pseudotypes of N-ras caused neurological disorders in 30% of the infected animals. These results differed markedly from those of previous experiments in which the authors had inserted the activated form of N-ras in the pSV(X) vector: the resulting SVN-ras virus was transforming on NIH 3T3 cells but was poorly oncogenic in vivo. Altogether, these data demonstrated unequivocally that N-ras is potentially as oncogenic as H-ras and that such oncogenic effect could depend on the vector environment.

  8. Live Imaging of Innate Immune and Preneoplastic Cell Interactions Using an Inducible Gal4/UAS Expression System in Larval Zebrafish Skin

    PubMed Central

    Ramezani, Thomas; Laux, Derek W.; Bravo, Isabel R.; Tada, Masazumi; Feng, Yi

    2015-01-01

    Here we describe a method to conditionally induce epithelial cell transformation by the use of the 4-Hydroxytamoxifen (4-OHT) inducible KalTA4-ERT2/UAS expression system1 in zebrafish larvae, and the subsequent live imaging of innate immune cell interaction with HRASG12V expressing skin cells. The KalTA4-ERT2/UAS system is both inducible and reversible which allows us to induce cell transformation with precise temporal/spatial resolution in vivo. This provides us with a unique opportunity to live image how individual preneoplastic cells interact with host tissues as soon as they emerge, then follow their progression as well as regression. Recent studies in zebrafish larvae have shown a trophic function of innate immunity in the earliest stages of tumorigenesis2,3. Our inducible system would allow us to live image the onset of cellular transformation and the subsequent host response, which may lead to important insights on the underlying mechanisms for the regulation of oncogenic trophic inflammatory responses. We also discuss how one might adapt our protocol to achieve temporal and spatial control of ectopic gene expression in any tissue of interest. PMID:25741625

  9. Common and overlapping oncogenic pathways contribute to the evolution of acute myeloid leukemias

    PubMed Central

    Kvinlaug, Brynn T; Chan, Wai-In; Bullinger, Lars; Ramaswami, Mukundhan; Sears, Christopher; Foster, Donna; Lazic, Stanley E; Okabe, Rachel; Benner, Axel; Lee, Benjamin H; De Silva, Inusha; Valk, Peter JM; Delwel, Ruud; Armstrong, Scott A; Döhner, Hartmut; Gilliland, D Gary; Huntly, Brian JP

    2011-01-01

    Fusion oncogenes in acute myeloid leukemia (AML) promote self-renewal from committed progenitors, thereby linking transformation and self-renewal pathways. Like most cancers, AML is a genetically and biologically heterogeneous disease, but it is unclear whether transformation results from common or overlapping genetic programs acting downstream of multiple mutations, or by the engagement of unique genetic programs acting cooperatively downstream of individual mutations. This distinction is important, because the involvement of common programs would imply the existence of common molecular targets to treat AML, no matter which fusion oncogenes are involved. Here we demonstrate that the ability to promote self-renewal is a generalized property of leukemia-associated oncogenes. Disparate oncogenes initiated overlapping transformation and self-renewal gene expression programs, the common elements of which were defined in established leukemia stem cells from an animal model as well as from a large cohort of patients with differing AML subtypes, where they strongly predicted pathobiological character. Notably, individual genes commonly activated in these programs could partially phenocopy the self-renewal function of leukemia-associated oncogenes in committed murine progenitors. Further, they could generate AML following expression in murine bone marrow. In summary, our findings reveal the operation of common programs of self-renewal and transformation downstream of leukemia-associated oncogenes, suggesting mechanistically common therapeutic approaches to AML are likely to be possible, regardless of the identity of the driver oncogene involved. PMID:21505102

  10. Serum screening for oncogene proteins in workers exposed to PCBs.

    PubMed Central

    Brandt-Rauf, P W; Niman, H L

    1988-01-01

    A cohort of 16 municipal workers engaged in cleaning oil from old transformers was examined for possible health effects from exposure to polychlorinated biphenyls (PCBs). In addition to the evaluation of routine clinical parameters (history, physical examination, liver function tests, serum triglycerides, serum PCB values), a new screening technique for the presence of oncogene proteins in serum using monoclonal antibodies was used to ascertain the potential carcinogenic risk from exposure in these workers. Except for one individual, serum PCB concentrations were found to be relatively low in this cohort, probably due to the observance of appropriate protective precautions. The results of liver function test were within normal limits and serum triglyceride concentrations showed no consistent relation to PCB concentrations. Six individuals, all of whom were smokers, showed abnormal banding patterns for fes oncogene related proteins. The individual with the highest serum PCB concentration also exhibited significantly raised levels of the H-ras oncogene related P21 protein in his serum. These oncogene protein findings may be indicative of an increased risk for the development of malignant disease in these individuals. Images PMID:3143397

  11. (Oncogenic action of ionizing radiation)

    SciTech Connect

    Not Available

    1990-01-01

    An extensive experiment involving approximately 400 rats exposed to the neon ion beam at the Bevalac in Berkeley, CA and to electrons is nearing completion. The carcinogenicity of energetic electrons was determined for comparison with the neon ion results. As in past reports we will describe progress in three areas corresponding to the specific aims of the proposal: (1) carcinogenesis and DNA strand breaks in rat skin following exposure by the neon ions or electrons; (2) DNA strand breaks in the epidermis as a function of radiation penetration; (3) oncogene activation in radiation-induced rat skin cancers. 72 refs., 6 tabs.

  12. Oncogenic Brain Metazoan Parasite Infection

    PubMed Central

    Spurgeon, Angela N.; Cress, Marshall C.; Gabor, Oroszi; Ding, Qing-Qing; Miller, Douglas C.

    2013-01-01

    Multiple observations suggest that certain parasitic infections can be oncogenic. Among these, neurocysticercosis is associated with increased risk for gliomas and hematologic malignancies. We report the case of a 71-year-old woman with colocalization of a metazoan parasite, possibly cysticercosis, and a WHO grade IV neuroepithelial tumor with exclusively neuronal differentiation by immunohistochemical stains (immunopositive for synaptophysin, neurofilament protein, and Neu-N and not for GFAP, vimentin, or S100). The colocalization and temporal relationship of these two entities suggest a causal relationship. PMID:24151568

  13. Oncogenic microtubule hyperacetylation through BEX4-mediated sirtuin 2 inhibition.

    PubMed

    Lee, Jin-Kwan; Lee, Janet; Go, Heounjeong; Lee, Chang Geun; Kim, Suhyeon; Kim, Hyun-Soo; Cho, Hyeseong; Choi, Kyeong Sook; Ha, Geun-Hyoung; Lee, Chang-Woo

    2016-01-01

    Five brain-expressed X-linked (BEX) gene members (BEX1-5) are arranged in tandem on chromosome X, and are highly conserved across diverse species. However, little is known about the function and role of BEX. This study represents a first attempt to demonstrate the molecular details of a novel oncogene BEX4. Among BEX proteins, BEX4 localizes to microtubules and spindle poles, and interacts with α-tubulin (α-TUB) and sirtuin 2 (SIRT2). The overexpression of BEX4 leads to the hyperacetylation of α-TUB by inhibiting SIRT2-mediated deacetylation. Furthermore, we found BEX4 expression conferred resistance to apoptotic cell death but led to acquisition of aneuploidy, and also increased the proliferating potential and growth of tumors. These results suggest that BEX4 overexpression causes an imbalance between TUB acetylation and deacetylation by SIRT2 inhibition and induces oncogenic aneuploidy transformation. PMID:27512957

  14. Malignant transformation of diploid human fibroblasts by transfection of oncogenes

    SciTech Connect

    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.

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

    SciTech Connect

    Pauklin, Siim . E-mail: spauklin@ut.ee; Kristjuhan, Arnold; Maimets, Toivo; Jaks, Viljar

    2005-08-26

    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, {beta}-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.

  16. Atlas of protein expression: image capture, analysis, and design of terabyte image database

    NASA Astrophysics Data System (ADS)

    Wu, Jiahua; Maslen, Gareth; Warford, Anthony; Griffin, Gareth; Xie, Jane; Crowther, Sandra; McCafferty, John

    2006-03-01

    The activity of genes in health and disease are manifested through the proteins which they encode. Ultimately, proteins drive functional processes in cells and tissues and so by measuring individual protein levels, studying modifications and discovering their sites of action we will understand better their function. It is possible to visualize the location of proteins of interest in tissue sections using labeled antibodies which bind to the target protein. This procedure, known as immunohistochemistry (IHC), provides valuable information on the cellular and sub-cellular distribution of proteins in tissue. The project, atlas of protein expression, aims to create a quality, information rich database of protein expression profiles, which is accessible to the world-wide research community. For the long term archival value of the data, the accompanying validated antibody and protein clones will potentially have great research, diagnostic and possibly therapeutic potential. To achieve this we had introduced a number of novel technologies, e.g. express recombinant proteins, select antibodies, stain proteins present in tissue section, and tissue microarray (TMA) image analysis. These are currently being optimized, automated and integrated into a multi-disciplinary production process. We had also created infrastructure for multi-terabyte scale image capture, established an image analysis capability for initial screening and quantization.

  17. Gender and Age Patterns in Emotional Expression, Body Image, and Self-Esteem: A Qualitative Analysis.

    ERIC Educational Resources Information Center

    Polce-Lynch, Mary; Myers, Barbara J.; Kilmartin, Christopher T.; Forssmann-Falck, Renate; Kliewer, Wendy

    1998-01-01

    Used written narratives to examine gender and age patterns in body image, emotional expression, and self-esteem for 209 students in grades 5, 8, and 12. Results indicate that boys restrict emotional expression in adolescence, whereas girls increase emotional expression in the same period. Girls also are more influenced by body image. (SLD)

  18. Comparison of liver oncogenic potential among human RAS isoforms

    PubMed Central

    Chung, Sook In; Moon, Hyuk; Ju, Hye-Lim; Kim, Dae Yeong; Cho, Kyung Joo; Ribback, Silvia; Dombrowski, Frank; Calvisi, Diego F.; Ro, Simon Weonsang

    2016-01-01

    Mutation in one of three RAS genes (i.e., HRAS, KRAS, and NRAS) leading to constitutive activation of RAS signaling pathways is considered a key oncogenic event in human carcinogenesis. Whether activated RAS isoforms possess different oncogenic potentials remains an unresolved question. Here, we compared oncogenic properties among RAS isoforms using liver-specific transgenesis in mice. Hydrodynamic transfection was performed using transposons expressing short hairpin RNA downregulating p53 and an activated RAS isoform, and livers were harvested at 23 days after gene delivery. No differences were found in the hepatocarcinogenic potential among RAS isoforms, as determined by both gross examination of livers and liver weight per body weight ratio (LW/BW) of mice expressing HRASQ61L, KRAS4BG12V and NRASQ61K. However, the tumorigenic potential differed significantly between KRAS splicing variants. The LW/BW ratio in KRAS4AG12V mice was significantly lower than in KRAS4BG12V mice (p < 0.001), and KRAS4AG12V mice lived significantly longer than KRRAS4BG12V mice (p < 0.0001). Notably, tumors from KRAS4AG12V mice displayed higher expression of the p16INK4A tumor suppressor when compared with KRAS4BG12V tumors. Forced overexpression of p16INK4A significantly reduced tumor growth in KRAS4BG12V mice, suggesting that upregulation of p16INK4A by KRAS4AG12V presumably delays tumor development driven by the latter oncogene. PMID:26799184

  19. High incidence of lung, bone, and lymphoid tumors in transgenic mice overexpressing mutant alleles of the p53 oncogene.

    PubMed Central

    Lavigueur, A; Maltby, V; Mock, D; Rossant, J; Pawson, T; Bernstein, A

    1989-01-01

    We have investigated the role of the p53 gene in oncogenesis in vivo by generating transgenic mice carrying murine p53 genomic fragments isolated from a mouse Friend erythroleukemia cell line or BALB/c mouse liver DNA. Elevated levels of p53 mRNA were detected in several tissues of two transgenic lines tested. Increased levels of p53 protein were also detected in most of the tissues analyzed by Western blotting (immunoblotting). Because both transgenes encoded p53 proteins that were antigenically distinct from wild-type p53, it was possible to demonstrate that overexpression of the p53 protein was mostly, if not entirely, due to the expression of the transgenes. Neoplasms developed in 20% of the transgenic mice, with a high incidence of lung adenocarcinomas, osteosarcomas, and lymphomas. Tissues such as ovaries that expressed the transgene at high levels were not at higher risk of malignant transformation than tissues expressing p53 protein at much lower levels. The long latent period and low penetrance suggest that overexpression of p53 alone is not sufficient to induce malignancies and that additional events are required. These observations provide direct evidence that mutant alleles of the p53 oncogene have oncogenic potential in vivo and that different cell types show intrinsic differences in susceptibility to malignant transformation by p53. Since recent data suggest that p53 may be a recessive oncogene, it is possible that the elevated tumor incidence results from functional inactivation of endogenous p53 by overexpression of the mutant transgene. The high incidence of lung and bone tumors suggests that p53 transgenic mice may provide a useful model to investigate the molecular events that underlie these malignancies in humans. Images PMID:2476668

  20. Functional imaging: monitoring heme oxygenase-1 gene expression in vivo

    NASA Astrophysics Data System (ADS)

    Zhang, Weisheng; Reilly-Contag, Pamela; Stevenson, David K.; Contag, Christopher H.

    1999-07-01

    The regulation of genetic elements can be monitored in living animals using photoproteins as reporters. Heme oxygenase (HO) is the key catabolic enzyme in the heme degradation pathway. Here, HO expression serves as a model for in vivo functional imaging of transcriptional regulation of a clinically relevant gene. HO enzymatic activity is inhibited by heme analogs, metalloporphyrins, but many members of this family of compounds also activate transcription of the HO-1 promoter. The degree of transcriptional activation by twelve metalloporphyrins, differing at the central metal and porphyrin ring substituents, was evaluated in both NIH 3T3 stable lines and transgenic animals containing HO-1 promoter-luciferase gene fusions. In the correlative cell culture assays, the metalloporphyrins increased transcription form the full length HO promoter fusion to varying degrees, but none increased transcription from a truncated HO-1 promoter. These results suggested that one or both of the two distal enhancer elements located at -4 and -10 Kb upstream from transcriptional start are required for HO-1 induction by heme and its analogs. The full-length HO-1-luc fusion was then evaluated as a transgene in mice. It was possible to monitor the effects of the metalloporphyrins, SnMP and ZnPP, in living animals over time. This spatiotemporal analyses of gene expression in vivo implied that alterations in porphyrin ring substituents and the central metal may affect the extent of gene activation. These data further indicate that using photoprotein reporters, subtle differences in gene expression can be monitored in living animals.

  1. Adenovirus E1A coding sequences that enable ras and pmt oncogenes to transform cultured primary cells.

    PubMed Central

    Zerler, B; Moran, B; Maruyama, K; Moomaw, J; Grodzicker, T; Ruley, H E

    1986-01-01

    Plasmids expressing partial adenovirus early region 1A (E1A) coding sequences were tested for activities which facilitate in vitro establishment (immortalization) of primary baby rat kidney cells and which enable the T24 Harvey ras-related oncogene and the polyomavirus middle T antigen (pmt) gene to transform primary baby rat kidney cells. E1A cDNAs expressing the 289- and 243-amino acid proteins expressed both E1A transforming functions. Mutant hrA, which encodes a 140-amino acid protein derived from the amino-terminal domain shared by the 289- and 243-amino acid proteins, enabled ras (but not pmt) to transform and facilitated in vitro establishment to a low, but detectable, extent. These studies suggest that E1A functions which collaborate with ras oncogenes and those which facilitate establishment are linked. Furthermore, E1A transforming functions are not associated with activities of the 289-amino acid E1A proteins required for efficient transcriptional activation of viral early region promoters. Images PMID:3022137

  2. Glucose metabolism and hexosamine pathway regulate oncogene-induced senescence.

    PubMed

    Gitenay, D; Wiel, C; Lallet-Daher, H; Vindrieux, D; Aubert, S; Payen, L; Simonnet, H; Bernard, D

    2014-01-01

    Oncogenic stress-induced senescence (OIS) prevents the ability of oncogenic signals to induce tumorigenesis. It is now largely admitted that the mitogenic effect of oncogenes requires metabolic adaptations to respond to new energetic and bio constituent needs. Yet, whether glucose metabolism affects OIS response is largely unknown. This is largely because of the fact that most of the OIS cellular models are cultivated in glucose excess. In this study, we used human epithelial cells, cultivated without glucose excess, to study alteration and functional role of glucose metabolism during OIS. We report a slowdown of glucose uptake and metabolism during OIS. Increasing glucose metabolism by expressing hexokinase2 (HK2), which converts glucose to glucose-6-phosphate (G6P), favors escape from OIS. Inversely, expressing a glucose-6-phosphatase, [corrected] pharmacological inhibition of HK2, or adding nonmetabolizable glucose induced a premature senescence. Manipulations of various metabolites covering G6P downstream pathways (hexosamine, glycolysis, and pentose phosphate pathways) suggest an unexpected role of the hexosamine pathway in controlling OIS. Altogether, our results show that decreased glucose metabolism occurs during and participates to OIS. PMID:24577087

  3. Utilizing signature-score to identify oncogenic pathways of cholangiocarcinoma

    PubMed Central

    Hsiao, Tzu-Hung; Chen, Hung-I Harry; Lu, Jo-Yang; Lin, Pei-Ying; Keller, Charles; Comerford, Sarah; Tomlinson, Gail E.; Chen, Yidong

    2013-01-01

    Extracting maximal information from gene signature sets (GSSs) via microarray-based transcriptional profiling involves assigning function to up and down regulated genes. Here we present a novel sample scoring method called Signature-score (S-score) which can be used to quantify the expression pattern of tumor samples from previously identified gene signature sets. A simulation result demonstrated an improved accuracy and robustness by S-score method comparing with other scoring methods. By applying the S-score method to cholangiocarcinoma (CAC), an aggressive hepatic cancer that arises from bile ducts cells, we identified enriched oncogenic pathways in two large CAC data sets. Thirteen pathways were enriched in CAC compared with normal liver and bile duct. Moreover, using S-score, we were able to dissect correlations between CAC-associated oncogenic pathways and Gene Ontology function. Two major oncogenic clusters and associated functions were identified. Cluster 1, which included beta-catenin and Ras, showed a positive correlation with the cell cycle, while cluster 2, which included TGF-beta, cytokeratin 19 and EpCAM was inversely correlated with immune function. We also used S-score to identify pathways that are differentially expressed in CAC and hepatocellular carcinoma (HCC), the more common subtype of liver cancer. Our results demonstrate the utility and effectiveness of S-score in assigning functional roles to tumor-associated gene signature sets and in identifying potential therapeutic targets for specific liver cancer subtypes. PMID:23905013

  4. SUMOylated IRF-1 shows oncogenic potential by mimicking IRF-2

    SciTech Connect

    Park, Sun-Mi; Chae, Myounghee; Kim, Bo-Kyoung; Seo, Taegun; Jang, Ik-Soon; Choi, Jong-Soon; Kim, Il-Chul; Lee, Je-Ho; Park, Junsoo

    2010-01-01

    Interferon regulatory factor-1 (IRF-1) is an interferon-induced transcriptional activator that suppresses tumors by impeding cell proliferation. Recently, we demonstrated that the level of SUMOylated IRF-1 is elevated in tumor cells, and that SUMOylation of IRF-1 attenuates its tumor-suppressive function. Here we report that SUMOylated IRF-1 mimics IRF-2, an antagonistic repressor, and shows oncogenic potential. To demonstrate the role of SUMOylated IRF-1 in tumorigenesis, we used SUMO-IRF-1 recombinant protein. Stable expression of SUMO-IRF-1 in NIH3T3 cells resulted in focus formation and anchorage-independent growth in soft agar. Inoculation of SUMO-IRF-1-transfected cells into athymic nude mice resulted in tumor formation and infiltration of adipose tissues. Finally, we demonstrated that SUMO-IRF-1 transforms NIH3T3 cells in a dose-dependent manner suggesting that SUMOylated IRF-1 may act as an oncogenic protein in tumor cells.

  5. Characterization and immunotherapeutic potential of a monoclonal antibody against a ras oncogene transformed cell line

    SciTech Connect

    Ames, R.S. Jr.

    1986-01-01

    Transformed cells express cell surface antigens not present, or present in diminished amounts on normal cells. Monoclonal antibodies can be used to identify and biochemically characterize tumor-associated antigens. Monoclonal antibody (MoAb) 45-2D9 was produced by immunization of BALB/c mice with a transformed cell line (45-2D9) induced by transfection of NIH 3T3 cells with a c-H-ras oncogene in DNA isolated from a human lung carcinoma. By immunoperoxidase staining, this antibody binds to the 45-342 cells as well as to the ras transformed primary and 3 secondary transfectants, including the one used to induce 45-342, but not to other ras transformed cell lines. Murine tumors as well as human fetal and most normal adult tissues are not stained. This antibody does bind to a variety of human tumors, including lung adenocarcinomas, as well as breast, colon and esophageal carcinomas. The ability of MoAb 45-2D9 to target ricin toxin A chain (RTA) and radio-isotopes to gp74 expressing cells was investigated. An immunotoxin generated by conjugating RTA to MoAb 45-2D9 inhibits protein and DNA synthesis by the 45-342 cells. Radiolabeled antibody specifically localizes to and can be used to image subcutaneous and pulmonary gp74 expressing tumors in nu/nu mice. Monoclonal antibodies against oncogene transformed cell lines may be useful for the detection and characterization of tumor-associated antigens as well as for the development of new tumor therapeutic and diagnostic reagents.

  6. IGF-Binding Protein 2 – Oncogene or Tumor Suppressor?

    PubMed Central

    Pickard, Adam; McCance, Dennis J.

    2015-01-01

    The role of insulin-like growth factor binding protein 2 (IGFBP2) in cancer is unclear. In general, IGFBP2 is considered to be oncogenic and its expression is often observed to be elevated in cancer. However, there are a number of conflicting reports in vitro and in vivo where IGFBP2 acts in a tumor suppressor manner. In this mini-review, we discuss the factors influencing the variation in IGFBP2 expression in cancer and our interpretation of these findings. PMID:25774149

  7. The human minisatellite consensus at breakpoints of oncogene translocations.

    PubMed Central

    Krowczynska, A M; Rudders, R A; Krontiris, T G

    1990-01-01

    A reexamination of human minisatellite (hypervariable) regions following the cloning and sequencing of the new minisatellite, VTR1.1, revealed that many of these structures possessed a strongly conserved copy of the chi-like octamer, GC[A/T]GG[A/T]GG. In oncogene translocations apparently created by aberrant VDJ recombinase activity, this VTR octamer was often found within a few bases of the breakpoint (p less than 10(-10)). Three bcl2 rearrangements which occurred within 2 bp of one another were located precisely adjacent to this consensus; it defined the 5' border of that oncogene's major breakpoint cluster. Several c-myc translocations also occurred within 2 bp of this sequence. While the appearance of a chi-like element in polymorphic minisatellite sequences is consistent with a role promoting either recombination or replication slippage, the existence of such elements at sites of somatic translocations suggests chi function in site-specific recombination, perhaps as a subsidiary recognition signal in immunoglobulin gene rearrangement. We discuss the implications of these observations for mechanisms by which oncogene translocations and minisatellite sequences are generated. Images PMID:1969618

  8. PERK Integrates Oncogenic Signaling and Cell Survival During Cancer Development.

    PubMed

    Bu, Yiwen; Diehl, J Alan

    2016-10-01

    Unfolded protein responses (UPR), consisting of three major transducers PERK, IRE1, and ATF6, occur in the midst of a variety of intracellular and extracellular challenges that perturb protein folding in the endoplasmic reticulum (ER). ER stress occurs and is thought to be a contributing factor to a number of human diseases, including cancer, neurodegenerative disorders, and various metabolic syndromes. In the context of neoplastic growth, oncogenic stress resulting from dysregulation of oncogenes such as c-Myc, Braf(V600E) , and HRAS(G12V) trigger the UPR as an adaptive strategy for cancer cell survival. PERK is an ER resident type I protein kinase harboring both pro-apoptotic and pro-survival capabilities. PERK, as a coordinator through its downstream substrates, reprograms cancer gene expression to facilitate survival in response to oncogenes and microenvironmental challenges, such as hypoxia, angiogenesis, and metastasis. Herein, we discuss how PERK kinase engages in tumor initiation, transformation, adaption microenvironmental stress, chemoresistance and potential opportunities, and potential opportunities for PERK targeted therapy. J. Cell. Physiol. 231: 2088-2096, 2016. © 2016 Wiley Periodicals, Inc. PMID:26864318

  9. CRAF R391W is a melanoma driver oncogene

    PubMed Central

    Atefi, Mohammad; Titz, Bjoern; Tsoi, Jennifer; Avramis, Earl; Le, Allison; Ng, Charles; Lomova, Anastasia; Lassen, Amanda; Friedman, Michael; Chmielowski, Bartosz; Ribas, Antoni; Graeber, Thomas G.

    2016-01-01

    Approximately 75% of melanomas have known driver oncogenic mutations in BRAF, NRAS, GNA11 or GNAQ, while the mutations providing constitutive oncogenic signaling in the remaining melanomas are not known. We established a melanoma cell line from a tumor with none of the common driver mutations. This cell line demonstrated a signaling profile similar to BRAF-mutants, but lacked sensitivity to the BRAF inhibitor vemurafenib. RNA-seq mutation data implicated CRAF R391W as the alternative driver mutation of this melanoma. CRAF R391W was homozygous and over expressed. These melanoma cells were highly sensitive to CRAF, but not BRAF knockdown. In reconstitution experiments, CRAF R391W, but not CRAF WT, transformed NIH3T3 cells in soft-agar colony formation assays, increased kinase activity in vitro, induced MAP kinase signaling and conferred vemurafenib resistance. MAP kinase inducing activity was dependent on CRAF dimerization. Thus, CRAF is a bona fide alternative oncogene for BRAF/NRAS/GNAQ/GNA11 wild type melanomas. PMID:27273450

  10. Oncogene-tumor suppressor gene feedback interactions and their control.

    PubMed

    Aguda, Baltazar D; del Rosario, Ricardo C H; Chan, Michael W Y

    2015-12-01

    We propose the hypothesis that for a particular type of cancer there exists a key pair of oncogene (OCG) and tumor suppressor gene (TSG) that is normally involved in strong stabilizing negative feedback loops (nFBLs) of molecular interactions, and it is these interactions that are sufficiently perturbed during cancer development. These nFBLs are thought to regulate oncogenic positive feedback loops (pFBLs) that are often required for the normal cellular functions of oncogenes. Examples given in this paper are the pairs of MYC and p53, KRAS and INK4A, and E2F1 and miR-17-92. We propose dynamical models of the aforementioned OCG-TSG interactions and derive stability conditions of the steady states in terms of strengths of cycles in the qualitative interaction network. Although these conditions are restricted to predictions of local stability, their simple linear expressions in terms of competing nFBLs and pFBLs make them intuitive and practical guides for experimentalists aiming to discover drug targets and stabilize cancer networks. PMID:26775863

  11. Oncogenic transformation of diverse gastrointestinal tissues in primary organoid culture

    PubMed Central

    Li, Xingnan; Nadauld, Lincoln; Ootani, Akifumi; Corney, David C.; Pai, Reetesh K.; Gevaert, Olivier; Cantrell, Michael A.; Rack, Paul G.; Neal, James T.; Chan, Carol W-M.; Yeung, Trevor; Gong, Xue; Yuan, Jenny; Wilhelmy, Julie; Robine, Sylvie; Attardi, Laura D.; Plevritis, Sylvia K.; Hung, Kenneth E.; Chen, Chang-Zheng; Ji, Hanlee P.; Kuo, Calvin J.

    2014-01-01

    The application of primary organoid cultures containing epithelial and mesenchymal elements to cancer modeling holds promise for combining the accurate multilineage differentiation and physiology of in vivo systems with the facile in vitro manipulation of transformed cell lines. Here, a single air-liquid interface culture method was used without modification to engineer oncogenic mutations into primary epithelial/mesenchymal organoids from mouse colon, stomach and pancreas. Pancreatic and gastric organoids exhibited dysplasia upon KrasG12D expression and/or p53 loss, and readily generated adenocarcinoma upon in vivo transplantation. In contrast, primary colon organoids required combinatorial Apc, p53, KrasG12D and Smad4 mutations for progressive transformation to invasive adenocarcinoma-like histology in vitro and tumorigenicity in vivo, recapitulating multi-hit models of colorectal cancer (CRC), and versus more promiscuous transformation of small intestinal organoids. Colon organoid culture functionally validated the microRNA miR-483 as a dominant driver oncogene at the Insulin-like growth factor-2 (IGF2) 11p15.5 CRC amplicon, inducing dysplasia in vitro and tumorigenicity in vivo. These studies demonstrate the general utility of a highly tractable primary organoid system for cancer modeling and driver oncogene validation in diverse gastrointestinal tissues. PMID:24859528

  12. Lung cancers unrelated to smoking: characterized by single oncogene addiction?

    PubMed

    Suda, Kenichi; Tomizawa, Kenji; Yatabe, Yasushi; Mitsudomi, Tetsuya

    2011-08-01

    Lung cancer is a major cause of cancer-related mortality worldwide. Currently, adenocarcinoma is its most common histological subtype in many countries. In contrast with small cell lung cancer or squamous cell carcinoma, lung adenocarcinoma often arises in never-smokers, especially in East Asian countries, as well as in smokers. Adenocarcinoma in never-smokers is associated with a lower incidence of genetic alterations (i.e., somatic mutations, loss of heterozygosity, and methylation) than in smokers. In addition, most adenocarcinomas in never-smokers harbor one of the proto-oncogene aberrations that occur in a mutually exclusive manner (EGFR mutation, KRAS mutation, HER2 mutations, or ALK translocation). It is of note that the proliferation and survival of lung cancer cells that harbor one of these oncogenic aberrations depend on the signaling from each aberrantly activated oncoprotein (oncogene addiction). Therefore, most adenocarcinomas in never-smokers can be effectively treated by molecularly targeted drugs that inhibit each oncoprotein. Moreover, from a pathological aspect, lung adenocarcinoma in never-smokers is characterized by terminal respiratory unit-type adenocarcinoma and a particular gene expression profile. Finally, epidemiological analyses have identified many candidate causes of lung cancer in never-smokers (genetic, environmental, and hormonal factors). The elucidation of the particular features of lung cancer unrelated to smoking and the development of new therapeutic modalities may reduce the mortality from lung cancers in the future. PMID:21655907

  13. Oncogene-mediated tumor transformation sensitizes cells to autophagy induction.

    PubMed

    Gargini, Ricardo; García-Escudero, Vega; Izquierdo, Marta; Wandosell, Francisco

    2016-06-01

    The process of tumorigenesis induces alterations in numerous cellular pathways including the main eukaryotic metabolic routes. It has been recently demonstrated that autophagy is part of the oncogene-induced senescence phenotype although its role in tumor establishment has not been completely clarified. In the present study, we showed that non‑transformed cells are sensitized to mitochondrial stress and autophagy induction when they are transformed by oncogenes such as c-Myc or Ras. We observed that overexpression of c-Myc or Ras increased AMP-activated protein kinase (AMPK) phosphorylation and the expression of p62, a known partner for degradation by autophagy. The activation of AMPK was found to favor the activation of FoxO3 which was prevented by the inhibition of AMPK. The transcriptional activation mediated by FoxO3 upregulated genes such as BNIP3 and LC3. Finally, the transformation by oncogenes such as c-Myc and Ras predisposes tumor cells to autophagy induction as a consequence of mitochondrial stress and impairs tumor growth in vitro and in vivo, which may have therapeutic implications. PMID:27035659

  14. Targeting oncogenic Ras signaling in hematologic malignancies

    PubMed Central

    Ward, Ashley F.; Braun, Benjamin S.

    2012-01-01

    Ras proteins are critical nodes in cellular signaling that integrate inputs from activated cell surface receptors and other stimuli to modulate cell fate through a complex network of effector pathways. Oncogenic RAS mutations are found in ∼ 25% of human cancers and are highly prevalent in hematopoietic malignancies. Because of their structural and biochemical properties, oncogenic Ras proteins are exceedingly difficult targets for rational drug discovery, and no mechanism-based therapies exist for cancers with RAS mutations. This article reviews the properties of normal and oncogenic Ras proteins, the prevalence and likely pathogenic role of NRAS, KRAS, and NF1 mutations in hematopoietic malignancies, relevant animal models of these cancers, and implications for drug discovery. Because hematologic malignancies are experimentally tractable, they are especially valuable platforms for addressing the fundamental question of how to reverse the adverse biochemical output of oncogenic Ras in cancer. PMID:22898602

  15. RAS oncogenes: weaving a tumorigenic web

    PubMed Central

    Pylayeva-Gupta, Yuliya; Grabocka, Elda; Bar-Sagi, Dafna

    2013-01-01

    RAS proteins are essential components of signalling pathways that emanate from cell surface receptors. Oncogenic activation of these proteins owing to missense mutations is frequently detected in several types of cancer. A wealth of biochemical and genetic studies indicates that RAS proteins control a complex molecular circuitry that consists of a wide array of interconnecting pathways. In this Review, we describe how RAS oncogenes exploit their extensive signalling reach to affect multiple cellular processes that drive tumorigenesis. PMID:21993244

  16. Oncogenes and inflammation rewire host energy metabolism in the tumor microenvironment

    PubMed Central

    Martinez-Outschoorn, Ubaldo E; Curry, Joseph M; Ko, Ying-Hui; Lin, Zhao; Tuluc, Madalina; Cognetti, David; Birbe, Ruth C; Pribitkin, Edmund; Bombonati, Alessandro; Pestell, Richard G; Howell, Anthony; Sotgia, Federica; Lisanti, Michael P

    2013-01-01

    Here, we developed a model system to evaluate the metabolic effects of oncogene(s) on the host microenvironment. A matched set of “normal” and oncogenically transformed epithelial cell lines were co-cultured with human fibroblasts, to determine the “bystander” effects of oncogenes on stromal cells. ROS production and glucose uptake were measured by FACS analysis. In addition, expression of a panel of metabolic protein biomarkers (Caveolin-1, MCT1, and MCT4) was analyzed in parallel. Interestingly, oncogene activation in cancer cells was sufficient to induce the metabolic reprogramming of cancer-associated fibroblasts toward glycolysis, via oxidative stress. Evidence for “metabolic symbiosis” between oxidative cancer cells and glycolytic fibroblasts was provided by MCT1/4 immunostaining. As such, oncogenes drive the establishment of a stromal-epithelial “lactate-shuttle”, to fuel the anabolic growth of cancer cells. Similar results were obtained with two divergent oncogenes (RAS and NFκB), indicating that ROS production and inflammation metabolically converge on the tumor stroma, driving glycolysis and upregulation of MCT4. These findings make stromal MCT4 an attractive target for new drug discovery, as MCT4 is a shared endpoint for the metabolic effects of many oncogenic stimuli. Thus, diverse oncogenes stimulate a common metabolic response in the tumor stroma. Conversely, we also show that fibroblasts protect cancer cells against oncogenic stress and senescence by reducing ROS production in tumor cells. Ras-transformed cells were also able to metabolically reprogram normal adjacent epithelia, indicating that cancer cells can use either fibroblasts or epithelial cells as “partners” for metabolic symbiosis. The antioxidant N-acetyl-cysteine (NAC) selectively halted mitochondrial biogenesis in Ras-transformed cells, but not in normal epithelia. NAC also blocked stromal induction of MCT4, indicating that NAC effectively functions as an “MCT4

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

    PubMed Central

    Kooistra, Susanne Marije; Nørgaard, Lise Christine Rudkjær; Lees, Michael James; Steinhauer, Cornelia; Johansen, Jens Vilstrup; Helin, Kristian

    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 fibroblasts. This screen led to the identification of miR-378a-5p and in addition several other miRNAs that have previously been shown to play a role in senescence. We show that ectopic expression of miR-378a-5p reduces the expression of several senescence markers, including p16INK4A and senescence-associated β-galactosidase. Moreover, cells with ectopic expression of miR-378a-5p retain proliferative capacity even in the presence of an activated Braf oncogene. Finally, we identified several miR-378a-5p targets in diploid fibroblasts that might explain the mechanism by which the microRNA can delay OIS. We speculate that miR-378a-5p might positively influence tumor formation by delaying OIS, which is consistent with a known pro-oncogenic function of this microRNA. PMID:24651706

  18. [Study of Her-2/neu oncogene in relation to prognosis of human breast cancer].

    PubMed

    Chen, R S

    1993-10-01

    A follow-up study of 143 cases of human breast cancer for over 5 years proved that Her-2/neu oncogene overexpression is much more common in the high risk group (patients died within 5 years) in comparison with the low risk group (patients survived over 5 years). The difference between these 2 groups was statistically significant. The Her-2/neu oncogene positive rate in infiltrative ductal carcinoma was 33.3%, the lower the differentiation, the higher the positive rate. Histological typing is also related to the positive rate, comedocarcinoma (intraductal carcinoma) expresses the highest positive rate while lobular carcinoma the lowest. Selection of fixation fluid and the mastering of diagnostic criteria are also important. In the author's opinion, only membrane staining in monoclonal antibody C-erbB-2 can be recognized as truly positive. In conclusion, Her-2/neu oncogene expression can be used as a supplemental marker when considering prognosis in breast cancer. PMID:7909501

  19. Oncogene Overdose: Too Much of a Bad Thing for Oncogene-Addicted Cancer Cells

    PubMed Central

    Amin, Amit Dipak; Rajan, Soumya S.; Groysman, Matthew J.; Pongtornpipat, Praechompoo; Schatz, Jonathan H.

    2015-01-01

    Acquired resistance to targeted inhibitors remains a major, and inevitable, obstacle in the treatment of oncogene-addicted cancers. Newer-generation inhibitors may help overcome resistance mutations, and inhibitor combinations can target parallel pathways, but durable benefit to patients remains elusive in most clinical scenarios. Now, recent studies suggest a third approach may be available in some cases—exploitation of oncogene overexpression that may arise to promote resistance. Here, we discuss the importance of maintaining oncogenic signaling at “just-right” levels in cells, with too much signaling, or oncogene overdose, being potentially as detrimental as too little. This is highlighted in particular by recent studies of mutant-BRAF in melanoma and the fusion kinase nucleophosmin–anaplastic lymphoma kinase (NPM–ALK) in anaplastic large cell lymphoma. Oncogene overdose may be exploitable to prolong tumor control through intermittent dosing in some cases, and studies of acute lymphoid leukemias suggest that it may be specifically pharmacologically inducible. PMID:26688666

  20. Bioinspired Nanocomplex for Spatiotemporal Imaging of Sequential mRNA Expression in Differentiating Neural Stem Cells

    PubMed Central

    2015-01-01

    Messenger RNA plays a pivotal role in regulating cellular activities. The expression dynamics of specific mRNA contains substantial information on the intracellular milieu. Unlike the imaging of stationary mRNAs, real-time intracellular imaging of the dynamics of mRNA expression is of great value for investigating mRNA biology and exploring specific cellular cascades. In addition to advanced imaging methods, timely extracellular stimulation is another key factor in regulating the mRNA expression repertoire. The integration of effective stimulation and imaging into a single robust system would significantly improve stimulation efficiency and imaging accuracy, producing fewer unwanted artifacts. In this study, we developed a multifunctional nanocomplex to enable self-activating and spatiotemporal imaging of the dynamics of mRNA sequential expression during the neural stem cell differentiation process. This nanocomplex showed improved enzymatic stability, fast recognition kinetics, and high specificity. With a mechanism regulated by endogenous cell machinery, this nanocomplex realized the successive stimulating motif release and the dynamic imaging of chronological mRNA expression during neural stem cell differentiation without the use of transgenetic manipulation. The dynamic imaging montage of mRNA expression ultimately facilitated genetic heterogeneity analysis. In vivo lateral ventricle injection of this nanocomplex enabled endogenous neural stem cell activation and labeling at their specific differentiation stages. This nanocomplex is highly amenable as an alternative tool to explore the dynamics of intricate mRNA activities in various physiological and pathological conditions. PMID:25494492

  1. In vivo imaging of inducible tyrosinase gene expression with an ultrasound array-based photoacoustic system

    NASA Astrophysics Data System (ADS)

    Harrison, Tyler; Paproski, Robert J.; Zemp, Roger J.

    2012-02-01

    Tyrosinase, a key enzyme in the production of melanin, has shown promise as a reporter of genetic activity. While green fluorescent protein has been used extensively in this capacity, it is limited in its ability to provide information deep in tissue at a reasonable resolution. As melanin is a strong absorber of light, it is possible to image gene expression using tyrosinase with photoacoustic imaging technologies, resulting in excellent resolutions at multiple-centimeter depths. While our previous work has focused on creating and imaging MCF-7 cells with doxycycline-controlled tyrosinase expression, we have now established the viability of these cells in a murine model. Using an array-based photoacoustic imaging system with 5 MHz center frequency, we capture interleaved ultrasound and photoacoustic images of tyrosinase-expressing MCF-7 tumors both in a tissue mimicking phantom, and in vivo. Images of both the tyrosinase-expressing tumor and a control tumor are presented as both coregistered ultrasound-photoacoustic B-scan images and 3-dimensional photoacoustic volumes created by mechanically scanning the transducer. We find that the tyrosinase-expressing tumor is visible with a signal level 12dB greater than that of the control tumor in vivo. Phantom studies with excised tumors show that the tyrosinase-expressing tumor is visible at depths in excess of 2cm, and have suggested that our imaging system is sensitive to a transfection rate of less than 1%.

  2. Transformation of human cells by oncogenic viruses supports permissiveness for parvovirus H-1 propagation.

    PubMed Central

    Faisst, S; Schlehofer, J R; zur Hausen, H

    1989-01-01

    Parvovirus H-1 has been shown to suppress spontaneous and chemically or virally induced tumorigenesis in hamsters. In human cell culture systems propagation of H-1 is restricted to transformed cells, which are killed by H-1 infection, in contrast to normal diploid cells, which are nonpermissive for H-1. By analyzing the permissiveness of a variety of human cells for H-1, it was determined that the majority of tested transformed or immortalized cells which were permissive for H-1 contained the DNA of oncogenic viruses (human papillomavirus, simian virus 40, adenovirus, hepatitis B virus, Epstein-Barr virus, and human T-cell lymphotropic virus type I). Of six transformed cell lines negative for persisting tumor virus DNA, only two were permissive for H-1, while two were semipermissive and two were nonpermissive. Thus, persistence and expression of tumor virus functions appears to promote full permissiveness for H-1 in human cells. However, neither expression of genes of specific viral genomes nor the transformed state of apparently virus-free cells alone was sufficient to render human cells permissive for H-1. Therefore, the effect of tumor virus functions on H-1 in transformed cells seems to be indirect, probably mediated by cellular factors which are induced or switched off during the transformation process. It appears that similar factors are induced or switched off by 5-azacytidine or calcium phosphate, both known inducers of cellular gene expression. Images PMID:2495371

  3. The protein encoded by the rolB plant oncogene hydrolyses indole glucosides.

    PubMed Central

    Estruch, J J; Schell, J; Spena, A

    1991-01-01

    The rolB gene of Agrobacterium rhizogenes, whose expression stimulates the formation of roots by transformed plant tissues and other growth alterations in transgenic plants, codes for a beta-glucosidase able to hydrolyse indole-beta-glucosides. Indeed, we show that extracts of bacteria and/or plant tissue expressing the rolB protein hydrolyse indoxyl-beta-glucoside (plant indican). Because of the structural similarity between indoxyl-beta-glucoside and indole-3-acetyl-beta-glucoside (IAA-beta-glucoside), we propose that the physiological and developmental alterations in transgenic plants expressing the rolB gene could be the result of an increased intracellular auxin activity caused by the release of active auxins from inactive beta-glucosides. Thus two of the oncogenes carried by the T-DNA of the plant pathogen Agrobacterium rhizogenes (rolB and rolC) perturb plant growth and development by coding for beta-glucosidases with distinct specificities. Whereas the rolC beta-glucosidase releases cytokinins from their glucoside conjugates, the rolB encoded protein hydrolyses indole-beta-glucosides. The combined action of these two genes therefore is expected to modulate the intracellular concentration of two of the main growth factors active in plants. Images PMID:1915286

  4. Oncogenic NRAS Primes Primary Acute Myeloid Leukemia Cells for Differentiation.

    PubMed

    Brendel, Cornelia; Teichler, Sabine; Millahn, Axel; Stiewe, Thorsten; Krause, Michael; Stabla, Kathleen; Ross, Petra; Huynh, Minh; Illmer, Thomas; Mernberger, Marco; Barckhausen, Christina; Neubauer, Andreas

    2015-01-01

    RAS mutations are frequently found among acute myeloid leukemia patients (AML), generating a constitutively active signaling protein changing cellular proliferation, differentiation and apoptosis. We have previously shown that treatment of AML patients with high-dose cytarabine is preferentially beneficial for those harboring oncogenic RAS. On the basis of a murine AML cell culture model, we ascribed this effect to a RAS-driven, p53-dependent induction of differentiation. Hence, in this study we sought to confirm the correlation between RAS status and differentiation of primary blasts obtained from AML patients. The gene expression signature of AML blasts with oncogenic NRAS indeed corresponded to a more mature profile compared to blasts with wildtype RAS, as demonstrated by gene set enrichment analysis (GSEA) and real-time PCR analysis of myeloid ecotropic viral integration site 1 homolog (MEIS1) in a unique cohort of AML patients. In addition, in vitro cell culture experiments with established cell lines and a second set of primary AML cells showed that oncogenic NRAS mutations predisposed cells to cytarabine (AraC) driven differentiation. Taken together, our findings show that AML with inv(16) and NRAS mutation have a differentiation gene signature, supporting the notion that NRAS mutation may predispose leukemic cells to AraC induced differentiation. We therefore suggest that promotion of differentiation pathways by specific genetic alterations could explain the superior treatment outcome after therapy in some AML patient subgroups. Whether a differentiation gene expression status may generally predict for a superior treatment outcome in AML needs to be addressed in future studies. PMID:25901794

  5. Metabolic alterations accompanying oncogene-induced senescence

    PubMed Central

    Aird, Katherine M; Zhang, Rugang

    2014-01-01

    Senescence is defined as a stable cell growth arrest. Oncogene-induced senescence (OIS) occurs in normal primary human cells after activation of an oncogene in the absence of other cooperating oncogenic stimuli. OIS is therefore considered a bona fide tumor suppression mechanism in vivo. Indeed, overcoming OIS-associated stable cell growth arrest can lead to tumorigenesis. Although cells that have undergone OIS do not replicate their DNA, they remain metabolically active. A number of recent studies report significant changes in cellular metabolism during OIS, including alterations in nucleotide, glucose, and mitochondrial metabolism and autophagy. These alterations may be necessary for stable senescence-associated cell growth arrest, and overcoming these shifts in metabolism may lead to tumorigenesis. This review highlights what is currently known about alterations in cellular metabolism during OIS and the implication of OIS-associated metabolic changes in cellular transformation and the development of cancer therapeutic strategies. PMID:27308349

  6. Epigenetic Pathways of Oncogenic Viruses: Therapeutic Promises.

    PubMed

    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. PMID:26754591

  7. The role of human cervical cancer oncogene in cancer progression.

    PubMed

    Li, Xin-Yu; Wang, Xin

    2015-01-01

    Human cervical cancer oncogene (HCCR) was identified by differential display RT-PCR by screened abnormally expressed genes in cervical human cancers. The overexpressed gene is not only identified in cervical tissues, but also in various human cancers as leukemia/lymphoma, breast, stomach, colon, liver, kidney and ovarian cancer. For its special sensitivities and specificities in human breast cancer and hepatocellular carcinoma, it is expected to be a new biomarker to replace or combine with the existing biomarkers in the diagnose. The HCCR manifests as a negative regulator of the p53 tumor suppressor gene, and its expression is regulated by the PI3K/Akt signaling pathway, modulated by TCF/β-catenin, it also participates in induction of the c-kit proto-oncogene, in activation of PKC and telomerase activities, but the accurate biochemical mechanisms of how HCCR contributes to the malignancies is still unknown. The aim of this review is to summarize the roles of HCCR in cancer progression and the molecular mechanisms involved. PMID:26309489

  8. ER functions of oncogenes and tumor suppressors: Modulators of intracellular Ca(2+) signaling.

    PubMed

    Bittremieux, Mart; Parys, Jan B; Pinton, Paolo; Bultynck, Geert

    2016-06-01

    Intracellular Ca(2+) signals that arise from the endoplasmic reticulum (ER), the major intracellular Ca(2+)-storage organelle, impact several mitochondrial functions and dictate cell survival and cell death processes. Furthermore, alterations in Ca(2+) signaling in cancer cells promote survival and establish a high tolerance towards cell stress and damage, so that the on-going oncogenic stress does not result in the activation of cell death. Over the last years, the mechanisms underlying these oncogenic alterations in Ca(2+) signaling have started to emerge. An important aspect of this is the identification of several major oncogenes, including Bcl-2, Bcl-XL, Mcl-1, PKB/Akt, and Ras, and tumor suppressors, such as p53, PTEN, PML, BRCA1, and Beclin 1, as direct and critical regulators of Ca(2+)-transport systems located at the ER membranes, including IP3 receptors and SERCA Ca(2+) pumps. In this way, these proteins execute part of their function by controlling the ER-mitochondrial Ca(2+) fluxes, favoring either survival (oncogenes) or cell death (tumor suppressors). Oncogenic mutations, gene deletions or amplifications alter the expression and/or function of these proteins, thereby changing the delicate balance between oncogenes and tumor suppressors, impacting oncogenesis and favoring malignant cell function and behavior. In this review, we provided an integrated overview of the impact of the major oncogenes and tumor suppressors, often altered in cancer cells, on Ca(2+) signaling from the ER Ca(2+) stores. This article is part of a Special Issue entitled: Calcium and Cell Fate. Guest Editors: Jacques Haiech, Claus Heizmann, Joachim Krebs, Thierry Capiod and Olivier Mignen. PMID:26772784

  9. Lymphomas that recur after MYC suppression continue to exhibit oncogene addiction

    PubMed Central

    Choi, Peter S.; van Riggelen, Jan; Gentles, Andrew J.; Bachireddy, Pavan; Rakhra, Kavya; Adam, Stacey J.; Plevritis, Sylvia K.; Felsher, Dean W.

    2011-01-01

    The suppression of oncogenic levels of MYC is sufficient to induce sustained tumor regression associated with proliferative arrest, differentiation, cellular senescence, and/or apoptosis, a phenomenon known as oncogene addiction. However, after prolonged inactivation of MYC in a conditional transgenic mouse model of Eμ-tTA/tetO-MYC T-cell acute lymphoblastic leukemia, some of the tumors recur, recapitulating what is frequently observed in human tumors in response to targeted therapies. Here we report that these recurring lymphomas express either transgenic or endogenous Myc, albeit in many cases at levels below those in the original tumor, suggesting that tumors continue to be addicted to MYC. Many of the recurring lymphomas (76%) harbored mutations in the tetracycline transactivator, resulting in expression of the MYC transgene even in the presence of doxycycline. Some of the remaining recurring tumors expressed high levels of endogenous Myc, which was associated with a genomic rearrangement of the endogenous Myc locus or activation of Notch1. By gene expression profiling, we confirmed that the primary and recurring tumors have highly similar transcriptomes. Importantly, shRNA-mediated suppression of the high levels of MYC in recurring tumors elicited both suppression of proliferation and increased apoptosis, confirming that these tumors remain oncogene addicted. These results suggest that tumors induced by MYC remain addicted to overexpression of this oncogene. PMID:21969595

  10. CONSISTENT ONCOGENE METHYLATION CHANGES IN EPITHELIAL CELLS CHEMICALLY TRANSFORMED IN VITRO

    EPA Science Inventory

    Many cancers occurring in humans and In animals are accompanied by alterations in oncogene DNA secluences, amplification, or changes in expression (1,2). In some cases the changes are quite specific and prevalent such as in Burkitts's lymphoma, pancreatic, and thyroid carcinoma (...

  11. Molecular cloning of an activated human oncogene, homologous to v-raf, from primary stomach cancer.

    PubMed Central

    Shimizu, K; Nakatsu, Y; Sekiguchi, M; Hokamura, K; Tanaka, K; Terada, M; Sugimura, T

    1985-01-01

    Transfection with high molecular weight DNA from a primary stomach cancer induced foci of transformed NIH 3T3 cells, and the transformed cells were tumorigenic in nude mice. By screening with a human Alu-family probe, we isolated the human DNA sequence from the secondary transformant cells. This transforming sequence encompasses about 60 kilobase pairs and is unrelated to known human transforming genes. Examination of homologies between this sequence and retroviral oncogenes revealed that the human transforming sequence is closely related to the v-raf oncogene of murine transforming retrovirus 3611-MSV. Images PMID:3862088

  12. Implication of oncogenic signaling pathways as a treatment strategy for neurodegenerative disorders - contemporary approaches.

    PubMed

    Sieradzki, Adrian; Yendluri, Bharat B; Palacios, Hector H; Parvathaneni, Kalpana; Reddy, V Prakash; Obrenovich, Mark E; Gąsiorowski, Kazimierz; Leszek, Jerzy; Aliev, Gjumrakch

    2011-03-01

    Recent evidence has associated the aberrant, proximal re-expression of various cell cycle control elements with neuronal cell vulnerability in Alzheimer's and Parkinson's diseases, as a common chronic neurodegeneration. This phenomenon associated with oncogenic transduction pathway activation has attracted the interest of scientists all over the world for a few years now. The purpose of this paper is to outline areas of research related to oncogenic factors or medicines in the context of potential applications for future treatment of the above mentioned chronic and, largely, incurable diseases. PMID:21222633

  13. Analysis of gene expression levels in individual bacterial cells without image segmentation

    SciTech Connect

    Kwak, In Hae; Son, Minjun; Hagen, Stephen J.

    2012-05-11

    Highlights: Black-Right-Pointing-Pointer We present a method for extracting gene expression data from images of bacterial cells. Black-Right-Pointing-Pointer The method does not employ cell segmentation and does not require high magnification. Black-Right-Pointing-Pointer Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. Black-Right-Pointing-Pointer We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

  14. Alpha-fetoprotein-targeted reporter gene expression imaging in hepatocellular carcinoma

    PubMed Central

    Kim, Kwang Il; Chung, Hye Kyung; Park, Ju Hui; Lee, Yong Jin; Kang, Joo Hyun

    2016-01-01

    Hepatocellular carcinoma (HCC) is one of the most common cancers in Eastern Asia, and its incidence is increasing globally. Numerous experimental models have been developed to better our understanding of the pathogenic mechanism of HCC and to evaluate novel therapeutic approaches. Molecular imaging is a convenient and up-to-date biomedical tool that enables the visualization, characterization and quantification of biologic processes in a living subject. Molecular imaging based on reporter gene expression, in particular, can elucidate tumor-specific events or processes by acquiring images of a reporter gene’s expression driven by tumor-specific enhancers/promoters. In this review, we discuss the advantages and disadvantages of various experimental HCC mouse models and we present in vivo images of tumor-specific reporter gene expression driven by an alpha-fetoprotein (AFP) enhancer/promoter system in a mouse model of HCC. The current mouse models of HCC development are established by xenograft, carcinogen induction and genetic engineering, representing the spectrum of tumor-inducing factors and tumor locations. The imaging analysis approach of reporter genes driven by AFP enhancer/promoter is presented for these different HCC mouse models. Such molecular imaging can provide longitudinal information about carcinogenesis and tumor progression. We expect that clinical application of AFP-targeted reporter gene expression imaging systems will be useful for the detection of AFP-expressing HCC tumors and screening of increased/decreased AFP levels due to disease or drug treatment. PMID:27468205

  15. Alpha-fetoprotein-targeted reporter gene expression imaging in hepatocellular carcinoma.

    PubMed

    Kim, Kwang Il; Chung, Hye Kyung; Park, Ju Hui; Lee, Yong Jin; Kang, Joo Hyun

    2016-07-21

    Hepatocellular carcinoma (HCC) is one of the most common cancers in Eastern Asia, and its incidence is increasing globally. Numerous experimental models have been developed to better our understanding of the pathogenic mechanism of HCC and to evaluate novel therapeutic approaches. Molecular imaging is a convenient and up-to-date biomedical tool that enables the visualization, characterization and quantification of biologic processes in a living subject. Molecular imaging based on reporter gene expression, in particular, can elucidate tumor-specific events or processes by acquiring images of a reporter gene's expression driven by tumor-specific enhancers/promoters. In this review, we discuss the advantages and disadvantages of various experimental HCC mouse models and we present in vivo images of tumor-specific reporter gene expression driven by an alpha-fetoprotein (AFP) enhancer/promoter system in a mouse model of HCC. The current mouse models of HCC development are established by xenograft, carcinogen induction and genetic engineering, representing the spectrum of tumor-inducing factors and tumor locations. The imaging analysis approach of reporter genes driven by AFP enhancer/promoter is presented for these different HCC mouse models. Such molecular imaging can provide longitudinal information about carcinogenesis and tumor progression. We expect that clinical application of AFP-targeted reporter gene expression imaging systems will be useful for the detection of AFP-expressing HCC tumors and screening of increased/decreased AFP levels due to disease or drug treatment. PMID:27468205

  16. Robotics and dynamic image analysis for studies of gene expression in plant tissues.

    PubMed

    Hernandez-Garcia, Carlos M; Chiera, Joseph M; Finer, John J

    2010-01-01

    Gene expression in plant tissues is typically studied by destructive extraction of compounds from plant tissues for in vitro analyses. The methods presented here utilize the green fluorescent protein (gfp) gene for continual monitoring of gene expression in the same pieces of tissues, over time. The gfp gene was placed under regulatory control of different promoters and introduced into lima bean cotyledonary tissues via particle bombardment. Cotyledons were then placed on a robotic image collection system, which consisted of a fluorescence dissecting microscope with a digital camera and a 2-dimensional robotics platform custom-designed to allow secure attachment of culture dishes. Images were collected from cotyledonary tissues every hour for 100 hours to generate expression profiles for each promoter. Each collected series of 100 images was first subjected to manual image alignment using ImageReady to make certain that GFP-expressing foci were consistently retained within selected fields of analysis. Specific regions of the series measuring 300 x 400 pixels, were then selected for further analysis to provide GFP Intensity measurements using ImageJ software. Batch images were separated into the red, green and blue channels and GFP-expressing areas were identified using the threshold feature of ImageJ. After subtracting the background fluorescence (subtraction of gray values of non-expressing pixels from every pixel) in the respective red and green channels, GFP intensity was calculated by multiplying the mean grayscale value per pixel by the total number of GFP-expressing pixels in each channel, and then adding those values for both the red and green channels. GFP Intensity values were collected for all 100 time points to yield expression profiles. Variations in GFP expression profiles resulted from differences in factors such as promoter strength, presence of a silencing suppressor, or nature of the promoter. In addition to quantification of GFP intensity, the

  17. Systematic RNA interference reveals that oncogenic KRAS-driven cancers require TBK1.

    PubMed

    Barbie, David A; Tamayo, Pablo; Boehm, Jesse S; Kim, So Young; Moody, Susan E; Dunn, Ian F; Schinzel, Anna C; Sandy, Peter; Meylan, Etienne; Scholl, Claudia; Fröhling, Stefan; Chan, Edmond M; Sos, Martin L; Michel, Kathrin; Mermel, Craig; Silver, Serena J; Weir, Barbara A; Reiling, Jan H; Sheng, Qing; Gupta, Piyush B; Wadlow, Raymond C; Le, Hanh; Hoersch, Sebastian; Wittner, Ben S; Ramaswamy, Sridhar; Livingston, David M; Sabatini, David M; Meyerson, Matthew; Thomas, Roman K; Lander, Eric S; Mesirov, Jill P; Root, David E; Gilliland, D Gary; Jacks, Tyler; Hahn, William C

    2009-11-01

    The proto-oncogene KRAS is mutated in a wide array of human cancers, most of which are aggressive and respond poorly to standard therapies. Although the identification of specific oncogenes has led to the development of clinically effective, molecularly targeted therapies in some cases, KRAS has remained refractory to this approach. A complementary strategy for targeting KRAS is to identify gene products that, when inhibited, result in cell death only in the presence of an oncogenic allele. Here we have used systematic RNA interference to detect synthetic lethal partners of oncogenic KRAS and found that the non-canonical IkappaB kinase TBK1 was selectively essential in cells that contain mutant KRAS. Suppression of TBK1 induced apoptosis specifically in human cancer cell lines that depend on oncogenic KRAS expression. In these cells, TBK1 activated NF-kappaB anti-apoptotic signals involving c-Rel and BCL-XL (also known as BCL2L1) that were essential for survival, providing mechanistic insights into this synthetic lethal interaction. These observations indicate that TBK1 and NF-kappaB signalling are essential in KRAS mutant tumours, and establish a general approach for the rational identification of co-dependent pathways in cancer. PMID:19847166

  18. Role of papillomavirus oncogenes in human cervical cancer: Transgenic animal studies

    SciTech Connect

    Griep, A.E.; Lambert, P.F.

    1994-05-01

    Human papillomaviruses are believed to be etiologic agents for the majority of human cervical carcinoma, a common cancer that is a leading cause of death by cancer among women worldwide. In cervical carcinoma, a subset of papillomaviral genes, namely E6 and E7, are expressed. In vitro tissue culture studies indicate that HPV E6 and E7 are oncogenes, and that their oncogenicity is due in part to their capacity to inactivate cellular tumor suppressor genes. The behavior of E6 and E7 in vitro and the genetic evidence from analysis of human cancers suggest that the E6 and E7 genes play a significant role in the development of cervical cancer. This hypothesis is now being tested using animal models. In this review, we summarize our current knowledge of the oncogenicity of papillomavirus genes that has been generated through their study in transgenic mice. 82 refs., 4 figs., 1 tab.

  19. Pancreatitis-induced Inflammation Contributes to Pancreatic Cancer by Inhibiting Oncogene-Induced Senescence

    PubMed Central

    Guerra, Carmen; Collado, Manuel; Navas, Carolina; Schuhmacher, Alberto J; Hernández-Porras, Isabel; Cañamero, Marta; Rodriguez-Justo, Manuel; Serrano, Manuel; Barbacid, Mariano

    2016-01-01

    Pancreatic acinar cells of adult mice (≥P60) are resistant to transformation by some of the most robust oncogenic insults including expression of K-Ras oncogenes and loss of p16Ink4a/p19Arf or Trp53 tumor suppressors. Yet, these acinar cells yield pancreatic intraepithelial neoplasias (mPanIN) and ductal adenocarcinomas (mPDAC) if exposed to limited bouts of non-acute pancreatitis, providing they harbor K-Ras oncogenes. Pancreatitis contributes to tumor progression by abrogating the senescence barrier characteristic of low-grade mPanINs. Attenuation of pancreatitis-induced inflammation also accelerates tissue repair and thwarts mPanIN expansion. Patients with chronic pancreatitis display senescent PanINs, if they have received anti-inflammatory drugs. These results put forward the concept that anti-inflammatory treatment of people diagnosed with pancreatitis may reduce their risk of developing PDAC. PMID:21665147

  20. Oncogenic Transcription Factors: Cornerstones of Inflammation-Linked Pancreatic Carcinogenesis

    PubMed Central

    Baumgart, Sandra; Ellenrieder, Volker; Fernandez-Zapico, Martin E.

    2012-01-01

    Transcription factors are proteins that regulate gene expression by modulating the synthesis of messenger RNA. Since this process is frequently one dominant control point in the production of many proteins, transcription factors represent the key regulators of numerous cellular functions, including proliferation, differentiation, and apoptosis. Pancreatic cancer progression is characterized by the activation of inflammatory signaling pathways converging on a limited set of transcription factors that fine-tune gene expression patterns contributing to the growth and maintenance of these tumors. Thus, strategies targeting these transcriptional networks activated in pancreatic cancer cells could block the effects of upstream inflammatory responses participating in pancreatic tumorigenesis. In this article we review this field of research and summarize current strategies to target oncogenic transcription factors and their activating signaling networks in the treatment of pancreatic cancer. PMID:21997559

  1. Human lung epithelial cells progressed to malignancy through specific oncogenic manipulations

    PubMed Central

    Sato, Mitsuo; Larsen, Jill E.; Lee, Woochang; Sun, Han; Shames, David S.; Dalvi, Maithili P.; Ramirez, Ruben D.; Tang, Hao; DiMaio, J. Michael; Gao, Boning; Xie, Yang; Wistuba, Ignacio I.; Gazdar, Adi F.; Shay, Jerry W.; Minna, John D.

    2013-01-01

    We used CDK4/hTERT-immortalized normal human bronchial epithelial cells (HBECs) from several individuals to study lung cancer pathogenesis by introducing combinations of common lung cancer oncogenic changes (p53, KRAS, MYC) and followed the stepwise transformation of HBECs to full malignancy. This model demonstrated that: 1) the combination of five genetic alterations (CDK4, hTERT, sh-p53, KRASV12, and c-MYC) is sufficient for full tumorigenic conversion of HBECs; 2) genetically-identical clones of transformed HBECs exhibit pronounced differences in tumor growth, histology, and differentiation; 3) HBECs from different individuals vary in their sensitivity to transformation by these oncogenic manipulations; 4) high levels of KRASV12 are required for full malignant transformation of HBECs, however prior loss of p53 function is required to prevent oncogene-induced senescence; 5) over-expression of c-MYC greatly enhances malignancy but only in the context of sh-p53+KRASV12; 6) growth of parental HBECs in serum-containing medium induces differentiation while growth of oncogenically manipulated HBECs in serum increases in vivo tumorigenicity, decreases tumor latency, produces more undifferentiated tumors, and induces epithelial-to-mesenchymal transition (EMT); 7) oncogenic transformation of HBECs leads to increased sensitivity to standard chemotherapy doublets; 8) an mRNA signature derived by comparing tumorigenic vs. non-tumorigenic clones was predictive of outcome in lung cancer patients. Collectively, our findings demonstrate this HBEC model system can be used to study the effect of oncogenic mutations, their expression levels, and serum-derived environmental effects in malignant transformation, while also providing clinically translatable applications such as development of prognostic signatures and drug response phenotypes. PMID:23449933

  2. INO80 governs superenhancer-mediated oncogenic transcription and tumor growth in melanoma.

    PubMed

    Zhou, Bingying; Wang, Li; Zhang, Shu; Bennett, Brian D; He, Fan; Zhang, Yan; Xiong, Chengliang; Han, Leng; Diao, Lixia; Li, Pishun; Fargo, David C; Cox, Adrienne D; Hu, Guang

    2016-06-15

    Superenhancers (SEs) are large genomic regions with a high density of enhancer marks. In cancer, SEs are found near oncogenes and dictate cancer gene expression. However, how oncogenic SEs are regulated remains poorly understood. Here, we show that INO80, a chromatin remodeling complex, is required for SE-mediated oncogenic transcription and tumor growth in melanoma. The expression of Ino80, the SWI/SNF ATPase, is elevated in melanoma cells and patient melanomas compared with normal melanocytes and benign nevi. Furthermore, Ino80 silencing selectively inhibits melanoma cell proliferation, anchorage-independent growth, tumorigenesis, and tumor maintenance in mouse xenografts. Mechanistically, Ino80 occupies >90% of SEs, and its occupancy is dependent on transcription factors such as MITF and Sox9. Ino80 binding reduces nucleosome occupancy and facilitates Mediator recruitment, thus promoting oncogenic transcription. Consistently, genes co-occupied by Ino80 and Med1 are selectively expressed in melanomas compared with melanocytes. Together, our results reveal an essential role of INO80-dependent chromatin remodeling in SE function and suggest a novel strategy for disrupting SEs in cancer treatment. PMID:27340176

  3. Multi-wavelength photoacoustic imaging of inducible tyrosinase reporter gene expression in xenograft tumors.

    PubMed

    Paproski, Robert J; Heinmiller, Andrew; Wachowicz, Keith; Zemp, Roger J

    2014-01-01

    Photoacoustic imaging is an emerging hybrid imaging technology capable of breaking through resolution limits of pure optical imaging technologies imposed by optical-scattering to provide fine-resolution optical contrast information in deep tissues. We demonstrate the ability of multi-wavelength photoacoustic imaging to estimate relative gene expression distributions using an inducible expression system and co-register images with hemoglobin oxygen saturation estimates and micro-ultrasound data. Tyrosinase, the rate-limiting enzyme in melanin production, is used as a reporter gene owing to its strong optical absorption and enzymatic amplification mechanism. Tetracycline-inducible melanin expression is turned on via doxycycline treatment in vivo. Serial multi-wavelength imaging reveals very low estimated melanin expression in tumors prior to doxycycline treatment or in tumors with no tyrosinase gene present, but strong signals after melanin induction in tumors tagged with the tyrosinase reporter. The combination of new inducible reporters and high-resolution photoacoustic and micro-ultrasound technology is poised to bring a new dimension to the study of gene expression in vivo. PMID:24936769

  4. A facial expression image database and norm for Asian population: a preliminary report

    NASA Astrophysics Data System (ADS)

    Chen, Chien-Chung; Cho, Shu-ling; Horszowska, Katarzyna; Chen, Mei-Yen; Wu, Chia-Ching; Chen, Hsueh-Chih; Yeh, Yi-Yu; Cheng, Chao-Min

    2009-01-01

    We collected 6604 images of 30 models in eight types of facial expression: happiness, anger, sadness, disgust, fear, surprise, contempt and neutral. Among them, 406 most representative images from 12 models were rated by more than 200 human raters for perceived emotion category and intensity. Such large number of emotion categories, models and raters is sufficient for most serious expression recognition research both in psychology and in computer science. All the models and raters are of Asian background. Hence, this database can also be used when the culture background is a concern. In addition, 43 landmarks each of the 291 rated frontal view images were identified and recorded. This information should facilitate feature based research of facial expression. Overall, the diversity in images and richness in information should make our database and norm useful for a wide range of research.

  5. Radiohybridization PET imaging of KRAS G12D mRNA expression in human pancreas cancer xenografts with [(64)Cu]DO3A-peptide nucleic acid-peptide nanoparticles.

    PubMed

    Chakrabarti, A; Zhang, K; Aruva, M R; Cardi, C A; Opitz, A W; Wagner, N J; Thakur, M L; Wickstrom, E

    2007-06-01

    There is a compelling need to image pancreas cancer at an early stage. Human pancreas cancer cells display elevated levels of KRAS protein due to high copy numbers of KRAS mRNA, and elevated levels of insulin-like growth factor 1 receptor (IGF1R) due to overexpression of IGF1R mRNA. Therefore we hypothesized that pancreas cancer could be detected in vivo with a single probe that targets both KRAS mRNA and IGF1R. Because positron emission tomography (PET) is a sensitive imaging technique, we designed a probe incorporating the positron-emitting nuclide (64)Cu. The KRAS-specific hybridization probe consisted of 1,4,7-tris(carboxymethylaza)cyclododecane-10-aza-acetyl (DO3A) on the N-terminus of a peptide nucleic acid (PNA) hybridization sequence (GCCATCAGCTCC) linked to a cyclized IGF1 peptide analog (d-Cys-Ser-Lys-Cys) on the C-terminus, for IGF1R-mediated endocytosis. A series of such KRAS radiohybridization probes with 0, 1, 2 or 3 mismatches to KRAS G12D mRNA, including exact matches to wild type KRAS mRNA and KRAS G12V mRNA, along with a double d(Ala) replacement IGF1 peptide control, were assembled by continuous solid phase synthesis. To test the hypothesis that KRAS-IGF1 dual probes could specifically image KRAS mRNA expression noninvasively in human IGF1R-overexpressing AsPC1 pancreas cancer xenografts in immunocompromised mice, [(64)Cu]PNA radiohybridization probes and controls were administered by tail vein. The [(64)Cu]KRAS-IGF1 radiohybridization probe yielded strong tumor contrast in PET images, 8.6 +/- 1.4-fold more intense in the center of human pancreas cancer xenografts than in the contralateral muscle at 4 h post-injection. Control experiments with single base KRASmismatches, an IGF1 peptide mismatch, and a breast cancer xenograft lacking KRAS activation yielded weak tumor contrast images. These experiments are consistent with our hypothesis for noninvasive PET imaging of KRAS oncogene expression in pancreas cancer xenografts. Imaging oncogene m

  6. Expression of San Andreas fault on Seasat radar image

    NASA Technical Reports Server (NTRS)

    Sabins, F. F., Jr.; Blom, R.; Elachi, C.

    1980-01-01

    A Seasat image (23.5 cm wavelength) of the Durmid Hills in southern California, the San Andreas Fault was analyzed. It is shown that a prominent southeast trending tonal lineament exists that is bright on the southwest side and dark on the northeast side. The cause of the contrasting signatures on opposite sides of the lineament was determined and the geologic signficance of the lineament was evaluated.

  7. Characterization and recognition of mixed emotional expressions in thermal face image

    NASA Astrophysics Data System (ADS)

    Saha, Priya; Bhattacharjee, Debotosh; De, Barin K.; Nasipuri, Mita

    2016-05-01

    Facial expressions in infrared imaging have been introduced to solve the problem of illumination, which is an integral constituent of visual imagery. The paper investigates facial skin temperature distribution on mixed thermal facial expressions of our created face database where six are basic expressions and rest 12 are a mixture of those basic expressions. Temperature analysis has been performed on three facial regions of interest (ROIs); periorbital, supraorbital and mouth. Temperature variability of the ROIs in different expressions has been measured using statistical parameters. The temperature variation measurement in ROIs of a particular expression corresponds to a vector, which is later used in recognition of mixed facial expressions. Investigations show that facial features in mixed facial expressions can be characterized by positive emotion induced facial features and negative emotion induced facial features. Supraorbital is a useful facial region that can differentiate basic expressions from mixed expressions. Analysis and interpretation of mixed expressions have been conducted with the help of box and whisker plot. Facial region containing mixture of two expressions is generally less temperature inducing than corresponding facial region containing basic expressions.

  8. Affibody-mediated PET imaging of HER3 expression in malignant tumours

    PubMed Central

    Rosestedt, Maria; Andersson, Ken G.; Mitran, Bogdan; Tolmachev, Vladimir; Löfblom, John; Orlova, Anna; Ståhl, Stefan

    2015-01-01

    Human epidermal growth factor receptor 3 (HER3) is involved in the progression of various cancers and in resistance to therapies targeting the HER family. In vivo imaging of HER3 expression would enable patient stratification for anti-HER3 immunotherapy. Key challenges with HER3-targeting are the relatively low expression in HER3-positive tumours and HER3 expression in normal tissues. The use of positron-emission tomography (PET) provides advantages of high resolution, sensitivity and quantification accuracy compared to SPECT. Affibody molecules, imaging probes based on a non-immunoglobulin scaffold, provide high imaging contrast shortly after injection. The aim of this study was to evaluate feasibility of PET imaging of HER3 expression using 68Ga-labeled affibody molecules. The anti-HER3 affibody molecule HEHEHE-Z08698-NOTA was successfully labelled with 68Ga with high yield, purity and stability. The agent bound specifically to HER3-expressing cancer cells in vitro and in vivo. At 3 h pi, uptake of 68Ga-HEHEHE-Z08698-NOTA was significantly higher in xenografts with high HER3 expression (BT474, BxPC-3) than in xenografts with low HER3 expression (A431). In xenografts with high expression, tumour-to-blood ratios were >20, tumour-to-muscle >15, and tumour-to-bone >7. HER3-positive xenografts were visualised using microPET 3 h pi. In conclusion, PET imaging of HER3 expression is feasible using 68Ga-HEHEHE-Z08698-NOTA shortly after administration. PMID:26477646

  9. KIT oncogene inhibition drives intratumoral macrophage M2 polarization.

    PubMed

    Cavnar, Michael J; Zeng, Shan; Kim, Teresa S; Sorenson, Eric C; Ocuin, Lee M; Balachandran, Vinod P; Seifert, Adrian M; Greer, Jonathan B; Popow, Rachel; Crawley, Megan H; Cohen, Noah A; Green, Benjamin L; Rossi, Ferdinand; Besmer, Peter; Antonescu, Cristina R; DeMatteo, Ronald P

    2013-12-16

    Tumor-associated macrophages (TAMs) are a major component of the cancer microenvironment. Modulation of TAMs is under intense investigation because they are thought to be nearly always of the M2 subtype, which supports tumor growth. Gastrointestinal stromal tumor (GIST) is the most common human sarcoma and typically results from an activating mutation in the KIT oncogene. Using a spontaneous mouse model of GIST and 57 freshly procured human GISTs, we discovered that TAMs displayed an M1-like phenotype and function at baseline. In both mice and humans, the KIT oncoprotein inhibitor imatinib polarized TAMs to become M2-like, a process which involved TAM interaction with apoptotic tumor cells leading to the induction of CCAAT/enhancer binding protein (C/EBP) transcription factors. In human GISTs that eventually developed resistance to imatinib, TAMs reverted to an M1-like phenotype and had a similar gene expression profile as TAMs from untreated human GISTs. Therefore, TAM polarization depends on tumor cell oncogene activity and has important implications for immunotherapeutic strategies in human cancers. PMID:24323358

  10. Oncogenic KRAS confers chemoresistance by upregulating NRF2

    PubMed Central

    Tao, Shasha; Wang, Shue; Moghaddam, Seyed Javad; Ooi, Aikseng; Chapman, Eli; Wong, Pak K.; Zhang, Donna D.

    2014-01-01

    Oncogenic KRAS mutations found in 20–30% of all non-small cell lung cancers (NSCLC) are associated with chemoresistance and poor prognosis. Here we demonstrate that activation of the cell protective stress response gene NRF2 by KRAS is responsible for its ability to promote drug resistance. RNAi-mediated silencing of NRF2 was sufficient to reverse resistance to cisplatin elicited by ectopic expression of oncogenic KRAS in NSCLC cells. Mechanistically, KRAS increased NRF2 gene transcription through a TPA response element (TRE) located in the NRF2 promoter. In a mouse model of mutant KrasG12D-induced lung cancer, we found that suppressing the NRF2 pathway with the chemical inhibitor brusatol enhanced the antitumor efficacy of cisplatin. Co-treatment reduced tumor burden and improved survival. Our findings illuminate the mechanistic details of KRAS-mediated drug resistance and provide a preclinical rationale to improve the management of lung tumors harboring KRAS mutations with NRF2 pathway inhibitors. PMID:25339352

  11. Insulator dysfunction and oncogene activation in IDH mutant gliomas.

    PubMed

    Flavahan, William A; Drier, Yotam; Liau, Brian B; Gillespie, Shawn M; Venteicher, Andrew S; Stemmer-Rachamimov, Anat O; Suvà, Mario L; Bernstein, Bradley E

    2016-01-01

    Gain-of-function IDH mutations are initiating events that define major clinical and prognostic classes of gliomas. Mutant IDH protein produces a new onco-metabolite, 2-hydroxyglutarate, which interferes with iron-dependent hydroxylases, including the TET family of 5'-methylcytosine hydroxylases. TET enzymes catalyse a key step in the removal of DNA methylation. IDH mutant gliomas thus manifest a CpG island methylator phenotype (G-CIMP), although the functional importance of this altered epigenetic state remains unclear. Here we show that human IDH mutant gliomas exhibit hypermethylation at cohesin and CCCTC-binding factor (CTCF)-binding sites, compromising binding of this methylation-sensitive insulator protein. Reduced CTCF binding is associated with loss of insulation between topological domains and aberrant gene activation. We specifically demonstrate that loss of CTCF at a domain boundary permits a constitutive enhancer to interact aberrantly with the receptor tyrosine kinase gene PDGFRA, a prominent glioma oncogene. Treatment of IDH mutant gliomaspheres with a demethylating agent partially restores insulator function and downregulates PDGFRA. Conversely, CRISPR-mediated disruption of the CTCF motif in IDH wild-type gliomaspheres upregulates PDGFRA and increases proliferation. Our study suggests that IDH mutations promote gliomagenesis by disrupting chromosomal topology and allowing aberrant regulatory interactions that induce oncogene expression. PMID:26700815

  12. Insulator dysfunction and oncogene activation in IDH mutant gliomas

    PubMed Central

    Flavahan, William A.; Drier, Yotam; Liau, Brian B.; Gillespie, Shawn M.; Venteicher, Andrew S.; Stemmer-Rachamimov, Anat O.; Suvà, Mario L.; Bernstein, Bradley E.

    2015-01-01

    Gain-of-function IDH mutations are initiating events that define major clinical and prognostic classes of gliomas1,2. Mutant IDH protein produces a novel onco-metabolite, 2-hydroxyglutarate (2-HG), that interferes with iron-dependent hydroxylases, including the TET family of 5′-methylcytosine hydroxylases3–7. TET enzymes catalyze a key step in the removal of DNA methylation8,9. IDH mutant gliomas thus manifest a CpG island methylator phenotype (G-CIMP)10,11, though the functional significance of this altered epigenetic state remains unclear. Here we show that IDH mutant gliomas exhibit hyper-methylation at CTCF binding sites, compromising binding of this methylation-sensitive insulator protein. Reduced CTCF binding is associated with loss of insulation between topological domains and aberrant gene activation. We specifically demonstrate that loss of CTCF at a domain boundary permits a constitutive enhancer to aberrantly interact with the receptor tyrosine kinase gene PDGFRA, a prominent glioma oncogene. Treatment of IDH mutant gliomaspheres with demethylating agent partially restores insulator function and down-regulates PDGFRA. Conversely, CRISPR-mediated disruption of the CTCF motif in IDH wildtype gliomaspheres up-regulates PDGFRA and increases proliferation. Our study suggests that IDH mutations promote gliomagenesis by disrupting chromosomal topology and allowing aberrant regulatory interactions that induce oncogene expression. PMID:26700815

  13. Expression-invariant three-dimensional face reconstruction from a single image by facial expression generic elastic models

    NASA Astrophysics Data System (ADS)

    Moeini, Ali; Faez, Karim; Moeini, Hossein

    2014-09-01

    An efficient method for expression-invariant three-dimensional (3-D) face reconstruction from a frontal face image with a variety of facial expressions (FE) using the FE generic elastic model (GEM) is proposed. Three generic models are employed for FE modeling in the generic elastic model (GEM) framework, which are combined based on the similarity distance around the lips. Exclusively, FE-GEM demonstrated that it is more precisely able to estimate a 3-D model of a frontal face, attaining a more robust and better quality 3-D face reconstruction under a variety of FEs compared to the original GEM approach. It is tested on an available 3-D face database and its accuracy and robustness are demonstrated compared to the GEM approach under a variety of FEs. Also, the FE-GEM method is tested on available two-dimensional face databases and a new synthesized pose is generated from gallery images for handling pose variations in face recognition.

  14. Learning-based Segmentation Framework for Tissue Images Containing Gene Expression Data

    SciTech Connect

    Bello, Musodiq; Ju, Tao; Carson, James P.; Warren, Joe; Chiu, Wah; Kakadiaris, Ioannis

    2007-05-01

    Abstract Associating specific gene activity with functional locations in the brain results in a greater understanding of the role of the gene. To perform such an association for the over 20,000 genes in the mammalian genome, reliable automated methods that characterize the distribution of gene expression in relation to a standard anatomical model are required. In this paper, we propose a new automatic method that results in the segmentation of gene expression images into distinct anatomical regions in which the expression can be quantified and compared with other images. Our contribution is a novel hybrid atlas that utilizes a statistical shape model based on a subdivision mesh, texture differentiation at region boundaries, and features of anatomical landmarks to delineate boundaries of anatomical regions in gene expression images. This atlas, which provides a common coordinate system for internal brain data, was trained on 36 images manually annotated by neuroanatomists and tested on 64 images. Our framework has achieved a mean overlap ratio of up to 91 § 7% in this challenging dataset. This tool for large-scale annotation will help scientists interpret gene expression patterns more efficiently.

  15. Modelling the perceptual similarity of facial expressions from image statistics and neural responses.

    PubMed

    Sormaz, Mladen; Watson, David M; Smith, William A P; Young, Andrew W; Andrews, Timothy J

    2016-04-01

    The ability to perceive facial expressions of emotion is essential for effective social communication. We investigated how the perception of facial expression emerges from the image properties that convey this important social signal, and how neural responses in face-selective brain regions might track these properties. To do this, we measured the perceptual similarity between expressions of basic emotions, and investigated how this is reflected in image measures and in the neural response of different face-selective regions. We show that the perceptual similarity of different facial expressions (fear, anger, disgust, sadness, happiness) can be predicted by both surface and feature shape information in the image. Using block design fMRI, we found that the perceptual similarity of expressions could also be predicted from the patterns of neural response in the face-selective posterior superior temporal sulcus (STS), but not in the fusiform face area (FFA). These results show that the perception of facial expression is dependent on the shape and surface properties of the image and on the activity of specific face-selective regions. PMID:26825440

  16. Facial expression recognition based on image Euclidean distance-supervised neighborhood preserving embedding

    NASA Astrophysics Data System (ADS)

    Chen, Li; Li, Yingjie; Li, Haibin

    2014-11-01

    High-dimensional data often lie on relatively low-dimensional manifold, while the nonlinear geometry of that manifold is often embedded in the similarities between the data points. These similar structures are captured by Neighborhood Preserving Embedding (NPE) effectively. But NPE as an unsupervised method can't utilize class information to guide the procedure of nonlinear dimensionality reduction. They ignore the geometrical structure information of local data points and the spatial information of pixels, which leads to the failure of classification. For this problem, a feature extraction method based on Image Euclidean Distance-Supervised NPE (IED-SNPE) is proposed, and is applied to facial expression recognition. Firstly, it employs Image Euclidean Distance (IED) to characterize the dissimilarity of data points. And then the neighborhood graph of the input data is constructed according to a certain kind of dissimilarity between data points. Finally, it fuses prior nonlinear facial expression manifold of facial expression images and class-label information to extract discriminative features for expression recognition. In the classification experiments on JAFFE facial expression database, IED-SNPE is used for feature extraction and compared with NPE, SNPE, and IED-NPE. The results reveal that IED-SNPE not only the local structure of expression manifold preserves well but also explicitly considers the spatial relationships among pixels in the images. So it excels NPE in feature extraction and is highly competitive with those well-known feature extraction methods.

  17. Function of oncogenes in cancer development: a changing paradigm

    PubMed Central

    Vicente-Dueñas, Carolina; Romero-Camarero, Isabel; Cobaleda, Cesar; Sánchez-García, Isidro

    2013-01-01

    Tumour-associated oncogenes induce unscheduled proliferation as well as genomic and chromosomal instability. According to current models, therapeutic strategies that block oncogene activity are likely to selectively target tumour cells. However, recent evidences have revealed that oncogenes are only essential for the proliferation of some specific tumour cell types, but not all. Indeed, the latest studies of the interactions between the oncogene and its target cell have shown that oncogenes contribute to cancer development not only by inducing proliferation but also by developmental reprogramming of the epigenome. This provides the first evidence that tumorigenesis can be initiated by stem cell reprogramming, and uncovers a new role for oncogenes in the origin of cancer. Here we analyse these evidences and propose an updated model of oncogene function that can explain the full range of genotype–phenotype associations found in human cancer. Finally, we discuss how this vision opens new avenues for developing novel anti-cancer interventions. PMID:23632857

  18. Targeting of the rasT24 oncogene to the proximal convoluted tubules in transgenic mice results in hyperplasia and polycystic kidneys.

    PubMed Central

    Schaffner, D. L.; Barrios, R.; Massey, C.; Bañez, E. I.; Ou, C. N.; Rajagopalan, S.; Aguilar-Cordova, E.; Lebovitz, R. M.; Overbeek, P. A.; Lieberman, M. W.

    1993-01-01

    Five families of transgenic mice were derived from one-cell-stage embryos injected with gamma GT-rasT24, a fusion gene consisting of the gamma-glutamyl transpeptidase (gamma GT) 5' flanking region containing promoter I linked to a mutated (codon 12) human H-ras oncogene. The transgene was expressed selectively in the kidneys, eyes, and brains of all families as determined by reverse transcription-polymerase chain reaction, nuclease protection assays, and in situ hybridization. In two of five families, kidney lesions consisting of proximal tubular hyperplasia, renal cysts, and microadenomas developed in male animals; males also expressed higher levels of gamma GT/rasT24 RNA. Early lesions consisted of proximal tubular hyperplasia as defined by alkaline phosphatase histochemistry, gamma GT immunohistochemistry, and electron microscopy and could be correlated with the presence of rasT24 RNA within the cystic proximal tubular epithelium by in situ hybridization. Advanced lesions also involved other segments of the nephron and consisted of cysts lined by a flattened unicellular layer of attenuated epithelium. No rasT24 could be identified within cystic lesions of the distal nephron and collecting tubules by in situ hybridization, and they most likely arise by external compression. Animals from the two transgenic strains exhibiting cystic lesions die of renal failure beginning at 8 months of age. No difference in cell-cycle parameters or DNA ploidy between transgenic and control kidneys was identified by flow cytometric analysis. No renal carcinomas developed. The primary renal effects of the H-rasT24 oncogene in this model system consist of proximal tubular hyperplasia and polycystic kidneys. This model appears to provide a useful in vivo system for the study of ras oncogene function and control of renal cell proliferation. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8097368

  19. Structure of mutant human oncogene protein determined

    SciTech Connect

    Baum, R.

    1989-01-16

    The protein encoded by a mutant human oncogene differs only slightly in structure from the native protein that initiates normal cell division, a finding that may complicate efforts to develop inhibitors of the mutant protein. Previously, the x-ray structure of the protein encoded by the normal c-Ha-ras gene, a protein believed to signal cells to start or stop dividing through its interaction with guanosine triphosphate (GTP), was reported. The structure of the protein encoded by a transforming c-Ha-ras oncogene, in which a valine codon replaces the normal glycine codon at position 12 in the gene, has now been determined. The differences in the structures of the mutant and normal proteins are located primarily in a loop that interacts with the /beta/-phosphate of a bound guanosine diphosphate (GDP) molecule.

  20. A novel dithiocarbamate derivative induces cell apoptosis through p53-dependent intrinsic pathway and suppresses the expression of the E6 oncogene of human papillomavirus 18 in HeLa cells.

    PubMed

    Li, Yanhong; Qi, Hongxue; Li, Xiaobo; Hou, Xueling; Lu, Xueying; Xiao, Xiangwen

    2015-06-01

    Dithiocarbamates (DTCs) exhibit a broad spectrum of antitumor activities, however, their molecular mechanisms of antitumor have not yet been elucidated. Previously, we have synthesized a series of novel dithiocarbamate derivatives. These DTCs were examined for cytotoxic activities against five human cancer cell lines. In this study, one of dithiocarbamate (DTC1) with higher potential for HeLa cells was chosen to investigate molecular mechanisms for its anti-tumor activities. DTC1 could inhibit proliferation, and highly induce apoptosis in HeLa cells by activating caspase-3, -6 and -9; moreover, activities of caspase-3, -6 and -9 were inhibited by pan-caspase inhibitor, Z-VAD-FMK. Furthermore, DTC1 decreased the levels of Bcl-2 and Bcl-xL, and increased expression of cytosol cytochrome c, Bak, Bax and p53 in a time-dependent manner but had no effect on the level of Rb. It was shown that DTC1 induced HeLa cells apoptosis through a p53-dependent pathway as tested by the wild type p53 inhibitor, pifithrin-α. Additionally, the relative expression of E6 and E7 were evaluated in HPV18-positive (HeLa cells) by real-time PCR and western blotting. The results firstly demonstrated that DTC1 suppressed both expression of E6 mRNA and E6 oncoprotein, but had no effect on the expression of E7 mRNA and protein in HPV18. Our results suggested that DTC1 may serve as novel chemotherapeutic agents in the treatment of cervical cancer and potential anti-HPV virus candidates that merit further studies. PMID:25772545

  1. Ras-oncogenic Drosophila hindgut but not midgut cells use an inflammation-like program to disseminate to distant sites.

    PubMed

    Christofi, Theodoulakis; Apidianakis, Yiorgos

    2013-01-01

    The gastrointestinal tract is habitable by a variety of microorganisms and it is often a tissue inflicted by inflammation. Much discussion is raised in recent years about the role of microbiota in intestinal inflammation, but their role in intestinal cancer remains unclear. Here we discuss and extent our work on Drosophila melanogaster models of tumorigenesis and tumor cell invasion upon intestinal infection. In Drosophila midgut bacteria that cause enterocyte damage induce intestinal stem cell proliferation, which is diverted toward aberrant stem cell expansion upon oncogene expression to induce dysplastic tumors. In the hindgut though, oncogenes synergize with the innate immune response-not the bacterially mediated damage-to induce tumor cell invasion and dissemination to distant sites. Interestingly, our novel gene expression analysis of Drosophila hemocyte-like cells suggests commonalities with oncogenic hindgut cells in the innate immune response and the expression of matrix metalloproteinase 1 in response to bacterial infection. PMID:23060054

  2. Anti-tumor effects of genetic vaccines against HPV major oncogenes

    PubMed Central

    Cordeiro, Marcelo Nazário; Paolini, Francesca; Massa, Silvia; Curzio, Gianfranca; Illiano, Elena; Duarte Silva, Anna Jéssica; Franconi, Rosella; Bissa, Massimiliano; Morghen, Carlo De Giuli; de Freitas, Antonio Carlos; Venuti, Aldo

    2014-01-01

    Expression of HPV E5, E6 and E7 oncogenes are likely to overcome the regulation of cell proliferation and to escape immunological control, allowing uncontrolled growth and providing the potential for malignant transformation. Thus, their three oncogenic products may represent ideal target antigens for immunotherapeutic strategies. In previous attempts, we demonstrated that genetic vaccines against recombinant HPV16 E7 antigen were able to affect the tumor growth in a pre-clinical mouse model. To improve this anti-HPV strategy we developed a novel approach in which we explored the effects of E5-based genetic immunization. We designed novel HPV16 E5 genetic vaccines based on two different gene versions: whole E5 gene and E5Multi. The last one is a long multi epitope gene designed as a harmless E5 version. Both E5 genes were codon optimized for mammalian expression. In addition, we demonstrated that HPV 16 E5 oncogene is expressed in C3 mouse cell line making it an elective model for the study of E5 based vaccine. In this mouse model the immunological and biological activity of the E5 vaccines were assessed in parallel with the activity of anti-E7 and anti-E6 vaccines already reported to be effective in an immunotherapeutic setting. These E7 and E6 vaccines were made with mutated oncogenes, the E7GGG mutant that does not bind pRb and the E6F47R mutant that is less effective in inhibiting p53, respectively. Results confirmed the immunological activity of genetic formulations based on attenuated HPV16 oncogenes and showed that E5-based genetic immunization provided notable anti-tumor effects. PMID:25483514

  3. TARGETING ONCOGENIC BRAF IN HUMAN CANCER

    PubMed Central

    Pratilas, Christine; Xing, Feng; Solit, David

    2012-01-01

    MAPK pathway activation is a frequent event in human cancer and is often the result of activating mutations in the BRAF and RAS oncogenes. BRAF missense kinase domain mutations, the vast majority of which are V600E, occur in approximately 8% of human tumors. These mutations, which are non-overlapping in distribution with RAS mutations, are observed most frequently in melanoma but also in tumors arising in the colon, thyroid, lung and other sites. Supporting its classification as an oncogene, V600EBRAF stimulates ERK signaling, induces proliferation and is capable of promoting transformation. Given the frequent occurrence of BRAF mutations in human cancer and the continued requirement for BRAF activity in the tumors in which it is mutated, efforts are underway to develop targeted inhibitors of BRAF and its downstream effectors. These agents offer the possibility of greater efficacy and less toxicity than the systemic therapies currently available for tumors driven by activating mutations in the MAPK pathway. Early clinical results with the BRAF-selective inhibitors PLX4032 and GSK2118436 suggest that this strategy will prove successful in a select group of patients whose tumors are driven by oncogenic BRAF. PMID:21818706

  4. Oncogenes in Cell Survival and Cell Death

    PubMed Central

    Shortt, Jake; Johnstone, Ricky W.

    2012-01-01

    The transforming effects of proto-oncogenes such as MYC that mediate unrestrained cell proliferation are countered by “intrinsic tumor suppressor mechanisms” that most often trigger apoptosis. Therefore, cooperating genetic or epigenetic effects to suppress apoptosis (e.g., overexpression of BCL2) are required to enable the dual transforming processes of unbridled cell proliferation and robust suppression of apoptosis. Certain oncogenes such as BCR-ABL are capable of concomitantly mediating the inhibition of apoptosis and driving cell proliferation and therefore are less reliant on cooperating lesions for transformation. Accordingly, direct targeting of BCR-ABL through agents such as imatinib have profound antitumor effects. Other oncoproteins such as MYC rely on the anti-apoptotic effects of cooperating oncoproteins such as BCL2 to facilitate tumorigenesis. In these circumstances, where the primary oncogenic driver (e.g., MYC) cannot yet be therapeutically targeted, inhibition of the activity of the cooperating antiapoptotic protein (e.g., BCL2) can be exploited for therapeutic benefit. PMID:23209150

  5. Systematic processing of Mars Express HRSC panchromatic and colour image mosaics: Image equalisation using an external brightness reference

    NASA Astrophysics Data System (ADS)

    Michael, G. G.; Walter, S. H. G.; Kneissl, T.; Zuschneid, W.; Gross, C.; McGuire, P. C.; Dumke, A.; Schreiner, B.; van Gasselt, S.; Gwinner, K.; Jaumann, R.

    2016-02-01

    After more than ten years in orbit at Mars, the coverage from the High Resolution Stereo Camera (HRSC) on the European Space Agency's Mars Express is sufficient to begin constructing mosaic products on a global scale. We describe our systematic processing procedure and, in particular, the technique used to bring images affected by atmospheric dust into visual consistency with the mosaic. We outline how the same method is used to produce a relative colour mosaic which shows local colour differences. We demonstrate the results and show that the techniques may also be applied to images from other orbital cameras.

  6. Identification of Intrinsic Imaging Phenotypes for Breast Cancer Tumors: Preliminary Associations with Gene Expression Profiles1

    PubMed Central

    Ashraf, Ahmed Bilal; Daye, Dania; Gavenonis, Sara; Mies, Carolyn; Feldman, Michael; Rosen, Mark; Kontos, Despina

    2015-01-01

    Purpose To present a method for identifying intrinsic imaging phenotypes in breast cancer tumors and to investigate their association with prognostic gene expression profiles. Materials and Methods The authors retrospectively analyzed dynamic contrast material–enhanced (DCE) magnetic resonance (MR) images of the breast in 56 women (mean age, 55.6 years; age range, 37–74 years) diagnosed with estrogen receptor–positive breast cancer between 2005 and 2010. The study was approved by the institutional review board and compliant with HIPAA. The requirement to obtain informed consent was waived. Primary tumors were assayed with a validated gene expression assay that provides a score for the likelihood of recurrence. A multiparametric imaging phenotype vector was extracted for each tumor by using quantitative morphologic, kinetic, and spatial heterogeneity features. Multivariate linear regression was performed to test associations between DCE MR imaging features and recurrence likelihood. To identify intrinsic imaging phenotypes, hierarchical clustering was performed on the extracted feature vectors. Multivariate logistic regression was used to classify tumors at high versus low or medium risk of recurrence. To determine the additional value of intrinsic phenotypes, the phenotype category was tested as an additional variable. Receiver operating characteristic analysis and the area under the receiver operating characteristic curve (Az) were used to assess classification performance. Results There was a moderate correlation (r = 0.71, R2 = 0.50, P < .001) between DCE MR imaging features and the recurrence score. DCE MR imaging features were predictive of recurrence risk as determined by the surrogate assay, with an Az of 0.77 (P < .01). Four dominant imaging phenotypes were detected, with two including only low- and medium-risk tumors. When the phenotype category was used as an additional variable, the Az increased to 0.82 (P < .01). Conclusion Intrinsic imaging

  7. miR-454 functions as an oncogene by inhibiting CHD5 in hepatocellular carcinoma

    PubMed Central

    Sun, Lei; Yao, Hong; Lu, Baoling; Zhu, Liying

    2015-01-01

    Previous studies showed that miR-454 acted as an oncogene or tumor suppressor in cancer. However, its function in HCC remains unknown. In this study, we found that miR-454 expression was upregulated in HCC cell lines and tissues. Knockdown of miR-454 inhibited HCC cell proliferation and invasion and epithelial mesenchymal transition (EMT), whereas overexpression of miR-454 promoted HCC cell proliferation and invasion and EMT. Furthermore, we identified the CHD5 as a direct target of miR-454. CHD5 was downregulated in HCC tissues and cell lines and the expression level of CHD5 was inversely correlated with the expression of miR-454 in HCC tissues. In addition, knockdown of miR-454 inhibited the growth of HepG2-engrafted tumors in vivo. Taken together, these results indicated that miR-454 functioned as an oncogene in HCC. PMID:26287602

  8. In vivo imaging of clock gene expression in multiple tissues of freely moving mice.

    PubMed

    Hamada, Toshiyuki; Sutherland, Kenneth; Ishikawa, Masayori; Miyamoto, Naoki; Honma, Sato; Shirato, Hiroki; Honma, Ken-Ichi

    2016-01-01

    Clock genes are expressed throughout the body, although how they oscillate in unrestrained animals is not known. Here, we show an in vivo imaging technique that enables long-term simultaneous imaging of multiple tissues. We use dual-focal 3D tracking and signal-intensity calibration to follow gene expression in a target area. We measure circadian rhythms of clock genes in the olfactory bulb, right and left ears and cortices, and the skin. In addition, the kinetic relationship between gene expression and physiological responses to experimental cues is monitored. Under stable conditions gene expression is in phase in all tissues. In response to a long-duration light pulse, the olfactory bulb shifts faster than other tissues. In Cry1(-/-) Cry2(-/-) arrhythmic mice circadian oscillation is absent in all tissues. Thus, our system successfully tracks circadian rhythms in clock genes in multiple tissues in unrestrained mice. PMID:27285820

  9. The spit and image: a psychoanalytic dissection of a colloquial expression.

    PubMed

    Mahon, Eugene J

    2011-04-01

    "He is the spit of his father" or "he is the spit and image of his father" is a colloquial expression that has graced informal English for many centuries. When a "spitting image" made an entrance in the manifest content of an analysand's dream, it became possible to add a psychoanalytic point of view to an etymological and anthropological record. After discussing both this clinical case and an "anthropological case history," the author examines the subtle but complex genesis of this colloquial expression from a speculative applied psychoanalytic perspective. PMID:21627009

  10. Nuclear compartmentalization of the v-myb oncogene product.

    PubMed Central

    Boyle, W J; Lampert, M A; Li, A C; Baluda, M A

    1985-01-01

    Nuclei obtained from chicken leukemic myeloblasts transformed by avian myeloblastosis virus were fractionated into various subnuclear compartments, which were then analyzed by specific immunoprecipitation for the presence of the leukemogenic product, p48v-myb, of the viral oncogene. In cells labeled for 30 or 60 min with L-[35S]methionine and in unlabeled exponentially dividing leukemic cells analyzed by Western blotting, p48v-myb was detected within the nucleoplasm (29 +/- 9% [standard deviation] of the total), chromatin (7 +/- 4%), and lamina-nuclear matrix (64 +/- 9%). Also, in myeloblasts analyzed by immunofluorescence during mitosis, p48v-myb appeared to be dispersed through the cell like the lamina-nuclear matrix complex. Strong attachment to the nuclear matrix-lamina complex suggests that p48v-myb may be involved in DNA replication or transcription or both. Images PMID:3018495

  11. A Computational Drug Repositioning Approach for Targeting Oncogenic Transcription Factors.

    PubMed

    Gayvert, Kaitlyn M; Dardenne, Etienne; Cheung, Cynthia; Boland, Mary Regina; Lorberbaum, Tal; Wanjala, Jackline; Chen, Yu; Rubin, Mark A; Tatonetti, Nicholas P; Rickman, David S; Elemento, Olivier

    2016-06-14

    Mutations in transcription factor (TF) genes are frequently observed in tumors, often leading to aberrant transcriptional activity. Unfortunately, TFs are often considered undruggable due to the absence of targetable enzymatic activity. To address this problem, we developed CRAFTT, a computational drug-repositioning approach for targeting TF activity. CRAFTT combines ChIP-seq with drug-induced expression profiling to identify small molecules that can specifically perturb TF activity. Application to ENCODE ChIP-seq datasets revealed known drug-TF interactions, and a global drug-protein network analysis supported these predictions. Application of CRAFTT to ERG, a pro-invasive, frequently overexpressed oncogenic TF, predicted that dexamethasone would inhibit ERG activity. Dexamethasone significantly decreased cell invasion and migration in an ERG-dependent manner. Furthermore, analysis of electronic medical record data indicates a protective role for dexamethasone against prostate cancer. Altogether, our method provides a broadly applicable strategy for identifying drugs that specifically modulate TF activity. PMID:27264179

  12. A mouse model of melanoma driven by oncogenic KRAS

    PubMed Central

    Milagre, Carla; Dhomen, Nathalie; Geyer, Felipe C; Hayward, Robert; Lambros, Maryou; Reis-Filho, Jorge S; Marais, Richard

    2010-01-01

    The small G-protein NRAS is mutated in 22% of human melanomas, whereas the related proteins, KRAS and HRAS are mutated in only 2% and 1% of melanomas respectively. We have developed a mouse models of melanoma in which Cre recombinase/loxP technology is used to drive inducible expression of G12VKRAS in the melanocytic lineage. The mice develop skin hyper-pigmentation, nevi and tumors that bear many of the cardinal histopathology features and molecular characteristics of human melanoma. These tumors invade and destroy the underlying muscles and cells derived from them can grow as subcutaneous tumors and colonise the lungs of nude mice. These data establish that oncogenic KRAS can be a founder event in melanomagenesis. PMID:20516123

  13. The Activating Transcription Factor 3 Protein Suppresses the Oncogenic Function of Mutant p53 Proteins*

    PubMed Central

    Wei, Saisai; Wang, Hongbo; Lu, Chunwan; Malmut, Sarah; Zhang, Jianqiao; Ren, Shumei; Yu, Guohua; Wang, Wei; Tang, Dale D.; Yan, Chunhong

    2014-01-01

    Mutant p53 proteins (mutp53) often acquire oncogenic activities, conferring drug resistance and/or promoting cancer cell migration and invasion. Although it has been well established that such a gain of function is mainly achieved through interaction with transcriptional regulators, thereby modulating cancer-associated gene expression, how the mutp53 function is regulated remains elusive. Here we report that activating transcription factor 3 (ATF3) bound common mutp53 (e.g. R175H and R273H) and, subsequently, suppressed their oncogenic activities. ATF3 repressed mutp53-induced NFKB2 expression and sensitized R175H-expressing cancer cells to cisplatin and etoposide treatments. Moreover, ATF3 appeared to suppress R175H- and R273H-mediated cancer cell migration and invasion as a consequence of preventing the transcription factor p63 from inactivation by mutp53. Accordingly, ATF3 promoted the expression of the metastasis suppressor SHARP1 in mutp53-expressing cells. An ATF3 mutant devoid of the mutp53-binding domain failed to disrupt the mutp53-p63 binding and, thus, lost the activity to suppress mutp53-mediated migration, suggesting that ATF3 binds to mutp53 to suppress its oncogenic function. In line with these results, we found that down-regulation of ATF3 expression correlated with lymph node metastasis in TP53-mutated human lung cancer. We conclude that ATF3 can suppress mutp53 oncogenic function, thereby contributing to tumor suppression in TP53-mutated cancer. PMID:24554706

  14. Evidence that synthetic lethality underlies the mutual exclusivity of oncogenic KRAS and EGFR mutations in lung adenocarcinoma

    PubMed Central

    Unni, Arun M; Lockwood, William W; Zejnullahu, Kreshnik; Lee-Lin, Shih-Queen; Varmus, Harold

    2015-01-01

    Human lung adenocarcinomas (LUAD) contain mutations in EGFR in ∼15% of cases and in KRAS in ∼30%, yet no individual adenocarcinoma appears to carry activating mutations in both genes, a finding we have confirmed by re-analysis of data from over 600 LUAD. Here we provide evidence that co-occurrence of mutations in these two genes is deleterious. In transgenic mice programmed to express both mutant oncogenes in the lung epithelium, the resulting tumors express only one oncogene. We also show that forced expression of a second oncogene in human cancer cell lines with an endogenous mutated oncogene is deleterious. The most prominent features accompanying loss of cell viability were vacuolization, other changes in cell morphology, and increased macropinocytosis. Activation of ERK, p38 and JNK in the dying cells suggests that an overly active MAPK signaling pathway may mediate the phenotype. Together, our findings indicate that mutual exclusivity of oncogenic mutations may reveal unexpected vulnerabilities and therapeutic possibilities. DOI: http://dx.doi.org/10.7554/eLife.06907.001 PMID:26047463

  15. In vitro transformation of immature hematopoietic cells by the P210 BCR/ABL oncogene product of the Philadelphia chromosome

    SciTech Connect

    McLaughlin, J.; Chianese, E.; Witte, O.N.

    1987-09-01

    The Philadelphia chromosome is the cytogenetic hallmark of human chronic myelogenous leukemia. RNA splicing joins sequences from a gene on chromosome 22 (BCR) across the translocation breakpoint to a portion of the ABL oncogene from chromosome 9, resulting in a chimeric protein (P210) that is an active tyrosine kinase. Although strongly correlated with this specific human neoplasm, and implicated as an oncogene by analogy to the gene product of the Abelson murine leukemia virus, the P210 gene had not been tested directly for oncogenic potential in hematopoietic cells. The authors have used a retroviral gene-transfer system to express P210 in mouse bone marrow cells. When infected bone marrow is plated under conditions for long-term culture of cells of the B-lymphoid lineage, cells expressing high amounts of P210 tyrosine kinase dominate the culture and rapidly lead to clonal outgrowths of immature lymphoid cells. Expression of P210 is growth-stimulatory but not sufficient for full oncogenic behavior. Some clonal lines progress toward a fully malignant phenotype as judged by increased cloning efficiency in agar suspension and frequency and rapidity of tumor induction in syngeneic mice. Such in vitro systems should be useful in evaluating the sequential and perhaps synergistic involvement of the P210 gene and other oncogenes as models for the progressive changes observed in human chronic myelogenous leukemia.

  16. Intracranial phosphaturic mesenchymal tumor, mixed connective tissue variant presenting without oncogenic osteomalacia

    PubMed Central

    Bower, Regina S.; Daugherty, Wilson P.; Giannini, Caterina; Parney, Ian F.

    2012-01-01

    Background: Phosphaturic mesenchymal tumor, mixed connective tissue variant (PMTMCT) is a rare tumor typically occurring in soft tissues and bone, causing oncogenic (tumor-induced) osteomalacia (TIO) through secretion of the phosphaturic hormone, fibroblast growth factor-23 (FGF-23). Rare tumors identical to PMTMCT occur without known TIO. Intracranial localization of PMTMCT is extremely rare, with only two cases reported in the literature. We present a very unusual case of a patient with an intracranial PMTMCT that presented with neurologic changes without osteomalacia. Case Description: A 67-year-old woman presented with progressive incontinence, apathy, and abulia after having undergone a total knee replacement 1 month earlier. Imaging disclosed a large left frontal anterior fossa mass. She underwent uncomplicated surgical resection of this tumor. Surprisingly, histopathology suggested PMTMCT. Reverse transcription polymerase chain reaction (RT-PCR) assay demonstrating FGF-23 expression in the tumor confirmed the diagnosis. Serum FGF-23 levels postoperatively were normal and she had no clinical or laboratory evidence of osteomalacia or phosphaturia. Conclusion: This report should serve to alert clinicians to the possibility that PMTMCT can be included in the differential diagnosis of intracranial masses even in the absence of tumor-induced osteomalacia. PMID:23372968

  17. Decreased expression of hepatocyte nuclear factor 4α (Hnf4α)/microRNA-122 (miR-122) axis in hepatitis B virus-associated hepatocellular carcinoma enhances potential oncogenic GALNT10 protein activity.

    PubMed

    Wu, Qian; Liu, Hai-Ou; Liu, Yi-Dong; Liu, Wei-Si; Pan, Deng; Zhang, Wei-Juan; Yang, Liu; Fu, Qiang; Xu, Jie-Jie; Gu, Jian-Xin

    2015-01-01

    MicroRNA-122 (miR-122), a mammalian liver-specific miRNA, has been reported to play crucial roles in the control of diverse aspects of hepatic function and dysfunction, including viral infection and hepatocarcinogenesis. In this study, we explored the clinical significance, transcriptional regulation, and direct target of miR-122 in hepatitis B virus (HBV)-associated hepatocellular carcinoma. Reduced expression of miR-122 in patients with HBV-associated hepatocellular carcinoma was correlated with venous invasion and poor prognosis. Furthermore, UDP-N-acetyl-α-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase-10 (GALNT10) was identified as a bona fide target of miR-122 in hepatoma cells. Ectopic expression and knockdown studies showed that GALNT10 indeed promotes proliferation and apoptosis resistance of hepatoma cells in a glycosyltransferase-dependent manner. Critically, adverse correlation between miR-122 and GALNT10, a poor prognosticator of clinical outcome, was demonstrated in hepatoma patients. Hepatocyte nuclear factor 4α (Hnf4α), a liver-enriched transcription factor that activates miR-122 gene transcription, was suppressed in HBV-infected hepatoma cells. Chromatin immunoprecipitation assay showed significantly reduced association of Hnf4α with the miR-122 promoter in HBV-infected hepatoma cells. Moreover, GALNT10 was found to intensify O-glycosylation following signal activation of the epidermal growth factor receptor. In addition, in a therapeutic perspective, we proved that GALNT10 silencing increases sensitivity to sorafenib and doxorubicin challenge. In summary, our results reveal a novel Hnf4α/miR-122/GALNT10 regulatory pathway that facilitates EGF miR-122 activation and hepatoma growth in HBV-associated hepatocarcinogenesis. PMID:25422324

  18. Decreased Expression of Hepatocyte Nuclear Factor 4α (Hnf4α)/MicroRNA-122 (miR-122) Axis in Hepatitis B Virus-associated Hepatocellular Carcinoma Enhances Potential Oncogenic GALNT10 Protein Activity*

    PubMed Central

    Wu, Qian; Liu, Hai-Ou; Liu, Yi-Dong; Liu, Wei-Si; Pan, Deng; Zhang, Wei-Juan; Yang, Liu; Fu, Qiang; Xu, Jie-Jie; Gu, Jian-Xin

    2015-01-01

    MicroRNA-122 (miR-122), a mammalian liver-specific miRNA, has been reported to play crucial roles in the control of diverse aspects of hepatic function and dysfunction, including viral infection and hepatocarcinogenesis. In this study, we explored the clinical significance, transcriptional regulation, and direct target of miR-122 in hepatitis B virus (HBV)-associated hepatocellular carcinoma. Reduced expression of miR-122 in patients with HBV-associated hepatocellular carcinoma was correlated with venous invasion and poor prognosis. Furthermore, UDP-N-acetyl-α-d-galactosamine:polypeptide N-acetylgalactosaminyltransferase-10 (GALNT10) was identified as a bona fide target of miR-122 in hepatoma cells. Ectopic expression and knockdown studies showed that GALNT10 indeed promotes proliferation and apoptosis resistance of hepatoma cells in a glycosyltransferase-dependent manner. Critically, adverse correlation between miR-122 and GALNT10, a poor prognosticator of clinical outcome, was demonstrated in hepatoma patients. Hepatocyte nuclear factor 4α (Hnf4α), a liver-enriched transcription factor that activates miR-122 gene transcription, was suppressed in HBV-infected hepatoma cells. Chromatin immunoprecipitation assay showed significantly reduced association of Hnf4α with the miR-122 promoter in HBV-infected hepatoma cells. Moreover, GALNT10 was found to intensify O-glycosylation following signal activation of the epidermal growth factor receptor. In addition, in a therapeutic perspective, we proved that GALNT10 silencing increases sensitivity to sorafenib and doxorubicin challenge. In summary, our results reveal a novel Hnf4α/miR-122/GALNT10 regulatory pathway that facilitates EGF miR-122 activation and hepatoma growth in HBV-associated hepatocarcinogenesis. PMID:25422324

  19. Oncogenic Role of Merlin/NF2 in Glioblastoma

    PubMed Central

    Guerrero, Paola A.; Yin, Wei; Camacho, Laura; Marchetti, Dario

    2014-01-01

    Glioblastoma is the most common and aggressive primary brain tumor in adults, with a poor prognosis because of its resistance to radiotherapy and chemotherapy. Merlin/NF2 (neurofibromatosis type 2) is a tumor suppressor found to be mutated in most nervous system tumors; however, it is not mutated in glioblastomas. Merlin associates with several transmembrane receptors and intracellular proteins serving as an anchoring molecule. Additionally, it acts as a key component of cell motility. By selecting subpopulations of U251 glioblastoma cells, we observed that high expression of phosphorylated Merlin at serine 518 (S518-Merlin), Notch1 and epidermal growth factor receptor (EGFR) correlated with increased cell proliferation and tumorigenesis. These cells were defective in cell-contact inhibition with changes in Merlin phosphorylation directly affecting Notch1, EGFR expression as well as downstream targets Hes1 and Ccnd. Of note, we identified a function for S518-Merlin which is distinct from what has been reported when the expression of Merlin is diminished in relation to EGFR and Notch expression, providing first-time evidence that demonstrates that the phosphorylation of Merlin at S518 in glioblastoma promotes oncogenic properties that are not only the result of inactivation of the tumor suppressor role of Merlin, but also, an independent process implicating a Merlin-driven regulation of Notch1 and EGFR. PMID:25043298

  20. Oncogenic potential diverge among human papillomavirus type 16 natural variants

    SciTech Connect

    Sichero, Laura; Simao Sobrinho, Joao; Lina Villa, Luisa

    2012-10-10

    We compared E6/E7 protein properties of three different HPV-16 variants: AA, E-P and E-350G. Primary human foreskin keratinocytes (PHFK) were transduced with HPV-16 E6 and E7 and evaluated for proliferation and ability to grow in soft agar. E-P infected keratinocytes presented the lowest efficiency in colony formation. AA and E-350G keratinocytes attained higher capacity for in vitro transformation. We observed similar degradation of TP53 among HPV-16 variants. Furthermore, we accessed the expression profile in early (p5) and late passage (p30) transduced cells of 84 genes commonly involved in carcinogenesis. Most differences could be attributed to HPV-16 E6/E7 expression. In particular, we detected different expression of ITGA2 and CHEK2 in keratinocytes infected with AA and AA/E-350G late passage cells, respectively, and higher expression of MAP2K1 in E-350G transduced keratinocytes. Our results indicate differences among HPV-16 variants that could explain, at least in part, differences in oncogenic potential attributed to these variants.

  1. Oncogenic KRAS triggers MAPK-dependent errors in mitosis and MYC-dependent sensitivity to anti-mitotic agents.

    PubMed

    Perera, David; Venkitaraman, Ashok R

    2016-01-01

    Oncogenic KRAS induces cell proliferation and transformation, but little is known about its effects on cell division. Functional genetic screens have recently revealed that cancer cell lines expressing oncogenic KRAS are sensitive to interference with mitosis, but neither the mechanism nor the uniformity of anti-mitotic drug sensitivity connected with mutant KRAS expression are yet clear. Here, we report that acute expression of oncogenic KRAS in HeLa cells induces mitotic delay and defects in chromosome segregation through mitogen-activated protein kinase (MAPK) pathway activation and de-regulated expression of several mitosis-related genes. These anomalies are accompanied by increased sensitivity to anti-mitotic agents, a phenotype dependent on the transcription factor MYC and its downstream target anti-apoptotic protein BCL-XL. Unexpectedly, we find no correlation between KRAS mutational status or MYC expression levels and anti-mitotic drug sensitivity when surveying a large database of anti-cancer drug responses. However, we report that the co-existence of KRAS mutations and high MYC expression predicts anti-mitotic drug sensitivity. Our findings reveal a novel function of oncogenic KRAS in regulating accurate mitotic progression and suggest new avenues to therapeutically target KRAS-mutant tumours and stratify patients in ongoing clinical trials of anti-mitotic drugs. PMID:27412232

  2. Oncogenic KRAS triggers MAPK-dependent errors in mitosis and MYC-dependent sensitivity to anti-mitotic agents

    PubMed Central

    Perera, David; Venkitaraman, Ashok R.

    2016-01-01

    Oncogenic KRAS induces cell proliferation and transformation, but little is known about its effects on cell division. Functional genetic screens have recently revealed that cancer cell lines expressing oncogenic KRAS are sensitive to interference with mitosis, but neither the mechanism nor the uniformity of anti-mitotic drug sensitivity connected with mutant KRAS expression are yet clear. Here, we report that acute expression of oncogenic KRAS in HeLa cells induces mitotic delay and defects in chromosome segregation through mitogen-activated protein kinase (MAPK) pathway activation and de-regulated expression of several mitosis-related genes. These anomalies are accompanied by increased sensitivity to anti-mitotic agents, a phenotype dependent on the transcription factor MYC and its downstream target anti-apoptotic protein BCL-XL. Unexpectedly, we find no correlation between KRAS mutational status or MYC expression levels and anti-mitotic drug sensitivity when surveying a large database of anti-cancer drug responses. However, we report that the co-existence of KRAS mutations and high MYC expression predicts anti-mitotic drug sensitivity. Our findings reveal a novel function of oncogenic KRAS in regulating accurate mitotic progression and suggest new avenues to therapeutically target KRAS-mutant tumours and stratify patients in ongoing clinical trials of anti-mitotic drugs. PMID:27412232

  3. In-gel imaging of RNA processing using Broccoli reveals optimal aptamer expression strategies

    PubMed Central

    Filonov, Grigory S.; Kam, Christina W.; Song, Wenjiao; Jaffrey, Samie R.

    2015-01-01

    SUMMARY RNA aptamers can be expressed in cells to influence and image cellular processes. Aptamer folding is maintained by inserting the aptamers into highly structured RNA scaffolds. Here we show that commonly used RNA scaffolds exhibit unexpected instability and cleavage in bacterial and mammalian cells. Using an in-gel staining approach for rapid and simple detection of Spinach- or Broccoli-tagged RNAs in cells, we monitored the processing of RNAs tagged with scaffolded aptamers, revealing endonucleolytic cleavage, RNA instability and poor expression. We reengineered a natural three-way junction structure to generate an alternative scaffold that enables stable aptamer expression in cells. This scaffold was used to create cassettes containing up to four Broccoli units, markedly enhancing the brightness of mammalian cells expressing cassette-tagged RNAs. These experiments describe methods for screening RNA cleavage events in cells, and identify cell-compatible scaffolds that enable efficient tagging of RNAs with aptamers for cellular expression. PMID:26000751

  4. Identification of Tumor Suppressors and Oncogenes from Genomic and Epigenetic Features in Ovarian Cancer

    PubMed Central

    Wrzeszczynski, Kazimierz O.; Varadan, Vinay; Byrnes, James; Lum, Elena; Kamalakaran, Sitharthan; Levine, Douglas A.; Dimitrova, Nevenka; Zhang, Michael Q.; Lucito, Robert

    2011-01-01

    The identification of genetic and epigenetic alterations from primary tumor cells has become a common method to identify genes critical to the development and progression of cancer. We seek to identify those genetic and epigenetic aberrations that have the most impact on gene function within the tumor. First, we perform a bioinformatic analysis of copy number variation (CNV) and DNA methylation covering the genetic landscape of ovarian cancer tumor cells. We separately examined CNV and DNA methylation for 42 primary serous ovarian cancer samples using MOMA-ROMA assays and 379 tumor samples analyzed by The Cancer Genome Atlas. We have identified 346 genes with significant deletions or amplifications among the tumor samples. Utilizing associated gene expression data we predict 156 genes with altered copy number and correlated changes in expression. Among these genes CCNE1, POP4, UQCRB, PHF20L1 and C19orf2 were identified within both data sets. We were specifically interested in copy number variation as our base genomic property in the prediction of tumor suppressors and oncogenes in the altered ovarian tumor. We therefore identify changes in DNA methylation and expression for all amplified and deleted genes. We statistically define tumor suppressor and oncogenic features for these modalities and perform a correlation analysis with expression. We predicted 611 potential oncogenes and tumor suppressors candidates by integrating these data types. Genes with a strong correlation for methylation dependent expression changes exhibited at varying copy number aberrations include CDCA8, ATAD2, CDKN2A, RAB25, AURKA, BOP1 and EIF2C3. We provide copy number variation and DNA methylation analysis for over 11,500 individual genes covering the genetic landscape of ovarian cancer tumors. We show the extent of genomic and epigenetic alterations for known tumor suppressors and oncogenes and also use these defined features to identify potential ovarian cancer gene candidates. PMID

  5. Activation of proto-oncogenes by disruption of chromosome neighborhoods.

    PubMed

    Hnisz, Denes; Weintraub, Abraham S; Day, Daniel S; Valton, Anne-Laure; Bak, Rasmus O; Li, Charles H; Goldmann, Johanna; Lajoie, Bryan R; Fan, Zi Peng; Sigova, Alla A; Reddy, Jessica; Borges-Rivera, Diego; Lee, Tong Ihn; Jaenisch, Rudolf; Porteus, Matthew H; Dekker, Job; Young, Richard A

    2016-03-25

    Oncogenes are activated through well-known chromosomal alterations such as gene fusion, translocation, and focal amplification. In light of recent evidence that the control of key genes depends on chromosome structures called insulated neighborhoods, we investigated whether proto-oncogenes occur within these structures and whether oncogene activation can occur via disruption of insulated neighborhood boundaries in cancer cells. We mapped insulated neighborhoods in T cell acute lymphoblastic leukemia (T-ALL) and found that tumor cell genomes contain recurrent microdeletions that eliminate the boundary sites of insulated neighborhoods containing prominent T-ALL proto-oncogenes. Perturbation of such boundaries in nonmalignant cells was sufficient to activate proto-oncogenes. Mutations affecting chromosome neighborhood boundaries were found in many types of cancer. Thus, oncogene activation can occur via genetic alterations that disrupt insulated neighborhoods in malignant cells. PMID:26940867

  6. Improved detection of differentially expressed genes in microarray experiments through multiple scanning and image integration

    PubMed Central

    Romualdi, Chiara; Trevisan, Silvia; Celegato, Barbara; Costa, Germano; Lanfranchi, Gerolamo

    2003-01-01

    The variability of results in microarray technology is in part due to the fact that independent scans of a single hybridised microarray give spot images that are not quite the same. To solve this problem and turn it to our advantage, we introduced the approach of multiple scanning and of image integration of microarrays. To this end, we have developed specific software that creates a virtual image that statistically summarises a series of consecutive scans of a microarray. We provide evidence that the use of multiple imaging (i) enhances the detection of differentially expressed genes; (ii) increases the image homogeneity; and (iii) reveals false-positive results such as differentially expressed genes that are detected by a single scan but not confirmed by successive scanning replicates. The increase in the final number of differentially expressed genes detected in a microarray experiment with this approach is remarkable; 50% more for microarrays hybridised with targets labelled by reverse transcriptase, and 200% more for microarrays developed with the tyramide signal amplification (TSA) technique. The results have been confirmed by semi-quantitative RT–PCR tests. PMID:14627839

  7. Photoacoustic imaging of gene expression using tyrosinase as a reporter gene

    NASA Astrophysics Data System (ADS)

    Paproski, Robert J.; Forbrich, Alexander; Harrison, Tyler; Hitt, Mary; Zemp, Roger J.

    2011-03-01

    Optical reporter genes, such as green fluorescence protein, are powerful research tools that allow visualization of gene expression. We have successfully used tyrosinase as a reporter gene for photoacoustic imaging. Tyrosinase is the key regulatory enzyme in the production of melanin which has a broad optical absorption spectrum. MCF-7 cells were stably transfected with tyrosinase under the control of an inducible promoter. For photoacoustic experiments, MCF-7 cells were resuspended at 108 cells/mL and injected in 700 μm (inner diameter) plastic tubing. Photoacoustic signal of MCF-7 cells expressing tyrosinase were >20-fold greater than those of untransfected MCF-7 cells. Photoacoustic signal of tyrosinaseexpressing MCF-7 cells were approximately 2-fold lesser and greater than those of blood at 576 and 650 nm, respectively, suggesting that photoacoustic signal from blood and tyrosinase-expressing cells can be separated by dualwavelength analysis. Photoacoustic signal from tyrosinase-expressing MCF-7 cells covered by chicken tissue could even be detected at a laser penetration depth of 4 cm, suggesting that tyrosinase can be used to image gene expression in relatively deep tissues. The current data suggests that tyrosinase is a strong reporter gene for photoacoustic imaging.

  8. Oncogenic RAS-induced senescence in human primary thyrocytes: molecular effectors and inflammatory secretome involved

    PubMed Central

    Vizioli, Maria Grazia; Santos, Joana; Pilotti, Silvana; Mazzoni, Mara; Anania, Maria Chiara; Miranda, Claudia; Pagliardini, Sonia; Pierotti, Marco A.

    2014-01-01

    Oncogene-induced senescence (OIS) is a robust and sustained antiproliferative response to oncogenic stress and constitutes an efficient barrier to tumour progression. We have recently proposed that OIS may be involved in the pathogenesis of thyroid carcinoma by restraining tumour progression as well as the transition of well differentiated to more aggressive variants. Here, an OIS inducible model was established and used for dissecting the molecular mechanisms and players regulating senescence in human primary thyrocytes. We show that oncogenic RAS induces senescence in thyrocytes as judged by changes in cell morphology, activation of p16INK4a and p53/p21CIP1 tumour suppressor pathways, senescence-associated β-galactosidase (SA-β-Gal) activity, and induction of proinflammatory components including IL-8 and its receptor CXCR2. Using RNA interference (RNAi) we demonstrate that p16INK4a is necessary for the onset of senescence in primary thyrocytes as its depletion rescues RAS-induced senescence. Furthermore, we found that IL-8/CXCR2 network reinforces the growth arrest triggered by oncogenic RAS, as its abrogation is enough to resume proliferation. Importantly, we observed that CXCR2 expression coexists with OIS markers in thyroid tumour samples, suggesting that CXCR2 contributes to senescence, thus limiting thyroid tumour progression. PMID:25268744

  9. Oncogenic transformation by vrel requires an amino-terminal activation domain

    SciTech Connect

    Kamens, J.; Brent, R. . Dept. of Molecular Biology); Richardson, P.; Gilmore, T. . Dept. of Biology); Mosialos, G. . Dept. of Chemistry)

    1990-06-01

    The mechanism by which the products of the v-{ital rel} oncogene, the corresponding c-{ital rel} proto-oncogene, and the related {ital dorsal} gene of {ital Drosophila melanogaster} exert their effects is not clear. The authors show that the v-{ital rel}, chicken c-{ital rel}, and {ital dorsal} proteins activated gene expression when fused to LexA sequences and bound to DNA upstream of target genes in {ital Saccharomyces cerevisiae}. They have defined two distinct activation regions in the c-{ital rel} protein. Region I, located in the amino-terminal half of {ital rel} and {ital dorsal} proteins, contains no stretches of glutamines, prolines, or acidic amino acids and therefore may be a novel activation domain. Lesions in the v-{ital rel} protein that diminished or abolished oncogenic transformation of avian spleen cells correspondingly affected transcription activation by region I. Region II, located in the carboxy terminus of the c-{ital rel} protein, is highly acidic. Region II is not present in the v-{ital rel} protein or in a transforming mutant derivative of the c-{ital rel} protein. The authors' results show that the oncogenicity of Rel proteins requires activation region I and suggest that the biological function of {ital rel} and {ital dorsal} proteins depends on transcription activation by this region.

  10. Netrin-1 exerts oncogenic activities through enhancing Yes-associated protein stability

    PubMed Central

    Qi, Qi; Li, Dean Y.; Luo, Hongbo R.; Guan, Kun-Liang; Ye, Keqiang

    2015-01-01

    Yes-associated protein (YAP), a transcription coactivator, is the major downstream effector of the Hippo pathway, which plays a critical role in organ size control and cancer development. However, how YAP is regulated by extracellular stimuli in tumorigenesis remains incompletely understood. Netrin-1, a laminin-related secreted protein, displays proto-oncogenic activity in cancers. Nonetheless, the downstream signaling mediating its oncogenic effects is not well defined. Here we show that netrin-1 via its transmembrane receptors, deleted in colorectal cancer and uncoordinated-5 homolog, up-regulates YAP expression, escalating YAP levels in the nucleus and promoting cancer cell proliferation and migration. Inactivating netrin-1, deleted in colorectal cancer, or uncoordinated-5 homolog B (UNC5B) decreases YAP protein levels, abrogating cancer cell progression by netrin-1, whereas knockdown of mammalian STE20-like protein kinase 1/2 (MST1/2) or large tumor suppressor kinase 1/2 (Lats1/2), two sets of upstream core kinases of the Hippo pathway, has no effect in blocking netrin-1–induced up-regulation of YAP. Netrin-1 stimulates phosphatase 1A to dephosphorylate YAP, which leads to decreased ubiquitination and degradation, enhancing YAP accumulation and signaling. Hence, our findings support that netrin-1 exerts oncogenic activity through YAP signaling, providing a mechanism coupling extracellular signals to the nuclear YAP oncogene. PMID:26039999

  11. Melanoma: oncogenic drivers and the immune system

    PubMed Central

    Karachaliou, Niki; Pilotto, Sara; Teixidó, Cristina; Viteri, Santiago; González-Cao, María; Riso, Aldo; Morales-Espinosa, Daniela; Molina, Miguel Angel; Chaib, Imane; Santarpia, Mariacarmela; Richardet, Eduardo; Bria, Emilio

    2015-01-01

    Advances and in-depth understanding of the biology of melanoma over the past 30 years have contributed to a change in the consideration of melanoma as one of the most therapy-resistant malignancies. The finding that oncogenic BRAF mutations drive tumor growth in up to 50% of melanomas led to a molecular therapy revolution for unresectable and metastatic disease. Moving beyond BRAF, inactivation of immune regulatory checkpoints that limit T cell responses to melanoma has provided targets for cancer immunotherapy. In this review, we discuss the molecular biology of melanoma and we focus on the recent advances of molecularly targeted and immunotherapeutic approaches. PMID:26605311

  12. Hedgehog Cholesterolysis: Specialized Gatekeeper to Oncogenic Signaling.

    PubMed

    Callahan, Brian P; Wang, Chunyu

    2015-01-01

    Discussions of therapeutic suppression of hedgehog (Hh) signaling almost exclusively focus on receptor antagonism; however, hedgehog's biosynthesis represents a unique and potentially targetable aspect of this oncogenic signaling pathway. Here, we review a key biosynthetic step called cholesterolysis from the perspectives of structure/function and small molecule inhibition. Cholesterolysis, also called cholesteroylation, generates cholesterol-modified Hh ligand via autoprocessing of a hedgehog precursor protein. Post-translational modification by cholesterol appears to be restricted to proteins in the hedgehog family. The transformation is essential for Hh biological activity and upstream of signaling events. Despite its decisive role in generating ligand, cholesterolysis remains conspicuously unexplored as a therapeutic target. PMID:26473928

  13. Repurposing a Prokaryotic Toxin-Antitoxin System for the Selective Killing of Oncogenically Stressed Human Cells.

    PubMed

    Preston, Mark A; Pimentel, Belén; Bermejo-Rodríguez, Camino; Dionne, Isabelle; Turnbull, Alice; de la Cueva-Méndez, Guillermo

    2016-07-15

    Prokaryotes express intracellular toxins that pass unnoticed to carrying cells until coexpressed antitoxin partners are degraded in response to stress. Although not evolved to function in eukaryotes, one of these toxins, Kid, induces apoptosis in mammalian cells, an effect that is neutralized by its cognate antitoxin, Kis. Here we engineered this toxin-antitoxin pair to create a synthetic system that becomes active in human cells suffering a specific oncogenic stress. Inspired by the way Kid becomes active in bacterial cells, we produced a Kis variant that is selectively degraded in human cells expressing oncoprotein E6. The resulting toxin-antitoxin system functions autonomously in human cells, distinguishing those that suffer the oncogenic insult, which are killed by Kid, from those that do not, which remain protected by Kis. Our results provide a framework for developing personalized anticancer strategies avoiding off-target effects, a challenge that has been hardly tractable by other means thus far. PMID:26230535

  14. Pose-variant facial expression recognition using an embedded image system

    NASA Astrophysics Data System (ADS)

    Song, Kai-Tai; Han, Meng-Ju; Chang, Shuo-Hung

    2008-12-01

    In recent years, one of the most attractive research areas in human-robot interaction is automated facial expression recognition. Through recognizing the facial expression, a pet robot can interact with human in a more natural manner. In this study, we focus on the facial pose-variant problem. A novel method is proposed in this paper to recognize pose-variant facial expressions. After locating the face position in an image frame, the active appearance model (AAM) is applied to track facial features. Fourteen feature points are extracted to represent the variation of facial expressions. The distance between feature points are defined as the feature values. These feature values are sent to a support vector machine (SVM) for facial expression determination. The pose-variant facial expression is classified into happiness, neutral, sadness, surprise or anger. Furthermore, in order to evaluate the performance for practical applications, this study also built a low resolution database (160x120 pixels) using a CMOS image sensor. Experimental results show that the recognition rate is 84% with the self-built database.

  15. Image enhancement of the Super Resolution Channel (SRC) of the Mars Express HRSC experiment

    NASA Astrophysics Data System (ADS)

    Michael, G.; Neukum, G.

    2009-04-01

    The SRC or Super Resolution Channel is the framing imaging subsystem of the HRSC experiment on the ESA Mars Express spacecraft, designed to show high-resolution detail within the broad swath of the HRSC push-broom camera. By December 2008, it had obtained about 7500 images. The quality of the SRC images proved lower than the design expectation: the thermal conditions of the camera in space caused a distortion of the optics, seen in images as blurring and ghosting. Analysis of star images demonstrated that the point-spread function was asymmetrical with offset secondary intensity peaks. Close to pericentre, where the spacecraft ground track velocity is around 3.5 km/s, an exposure time of 0.6 ms is needed to avoid motion-smear in the image. Operation of the camera has shown that the optimal trade-off between motion-smear and an acceptable SNR is found at considerably higher exposure times, around 5 ms. Trials at improving the images with respect to both the thermal distortion and the motion-smear were made using the Richardson-Lucy algorithm, an iterative procedure for recovering an image blurred by a known point spread function.

  16. Adaptive and Background-Aware GAL4 Expression Enhancement of Co-registered Confocal Microscopy Images.

    PubMed

    Trapp, Martin; Schulze, Florian; Novikov, Alexey A; Tirian, Laszlo; J Dickson, Barry; Bühler, Katja

    2016-04-01

    GAL4 gene expression imaging using confocal microscopy is a common and powerful technique used to study the nervous system of a model organism such as Drosophila melanogaster. Recent research projects focused on high throughput screenings of thousands of different driver lines, resulting in large image databases. The amount of data generated makes manual assessment tedious or even impossible. The first and most important step in any automatic image processing and data extraction pipeline is to enhance areas with relevant signal. However, data acquired via high throughput imaging tends to be less then ideal for this task, often showing high amounts of background signal. Furthermore, neuronal structures and in particular thin and elongated projections with a weak staining signal are easily lost. In this paper we present a method for enhancing the relevant signal by utilizing a Hessian-based filter to augment thin and weak tube-like structures in the image. To get optimal results, we present a novel adaptive background-aware enhancement filter parametrized with the local background intensity, which is estimated based on a common background model. We also integrate recent research on adaptive image enhancement into our approach, allowing us to propose an effective solution for known problems present in confocal microscopy images. We provide an evaluation based on annotated image data and compare our results against current state-of-the-art algorithms. The results show that our algorithm clearly outperforms the existing solutions. PMID:26743993

  17. Class I PI3K in oncogenic cellular transformation

    PubMed Central

    Zhao, Li; Vogt, Peter K.

    2009-01-01

    Class I phosphoinositide 3-kinase (PI3K) is a dimeric enzyme, consisting of a catalytic and a regulatory subunit. The catalytic subunit occurs in four isoforms designated as p110α, p110β, p110γ and p110δ. These combine with several regulatory subunits; for p110α, β and δ the standard regulatory subunit is p85, for p110γ it is p101. PI3Ks play important roles in human cancer. PIK3CA, the gene encoding p110α, is mutated frequently in common cancers, including carcinoma of the breast, prostate, colon and endometrium. Eighty percent of these mutations are represented by one of three amino acid substitutions in the helical or kinase domains of the enzyme. The mutant p110α shows a gain of function in enzymatic and signaling activity and is oncogenic in cell culture and in animal model systems. Structural and genetic data suggest that the mutations affect regulatory inter- and intramolecular interactions and support the conclusion that there are at least two molecular mechanisms for the gain-of-function in p110α. One of these mechanisms operates largely independently of binding to p85, the other abolishes the requirement for an interaction with Ras. The non-alpha isoforms of p110 do not show cancer-specific mutations. However, they are often differentially expressed in cancer and, in contrast to p110α, wild-type non-alpha isoforms of p110 are oncogenic when overexpressed in cell culture. The isoforms of p110 have become promising drug targets. Isoform-selective inhibitors have been identified. Inhibitors that target exclusively the cancer-specific mutants of p110α constitute an important goal and challenge for current drug development. PMID:18794883

  18. A humanized antibody for imaging immune checkpoint ligand PD-L1 expression in tumors

    PubMed Central

    Gabrielson, Matthew; Lisok, Ala; Wharram, Bryan; Sysa-Shah, Polina; Azad, Babak Behnam; Pomper, Martin G.; Nimmagadda, Sridhar

    2016-01-01

    Antibodies targeting the PD-1/PD-L1 immune checkpoint lead to tumor regression and improved survival in several cancers. PD-L1 expression in tumors may be predictive of response to checkpoint blockade therapy. Because tissue samples might not always be available to guide therapy, we developed and evaluated a humanized antibody for non-invasive imaging of PD-L1 expression in tumors. Radiolabeled [111In]PD-L1-mAb and near-infrared dye conjugated NIR-PD-L1-mAb imaging agents were developed using the mouse and human cross-reactive PD-L1 antibody MPDL3280A. We tested specificity of [111In]PD-L1-mAb and NIR-PD-L1-mAb in cell lines and in tumors with varying levels of PD-L1 expression. We performed SPECT/CT imaging, biodistribution and blocking studies in NSG mice bearing tumors with constitutive PD-L1 expression (CHO-PDL1) and in controls (CHO). Results were confirmed in triple negative breast cancer (TNBC) (MDAMB231 and SUM149) and non-small cell lung cancer (NSCLC) (H2444 and H1155) xenografts with varying levels of PD-L1 expression. There was specific binding of [111In]PD-L1-mAb and NIR-PD-L1-mAb to tumor cells in vitro, correlating with PD-L1 expression levels. In mice bearing subcutaneous and orthotopic tumors, there was specific and persistent high accumulation of signal intensity in PD-L1 positive tumors (CHO-PDL1, MDAMB231, H2444) but not in controls. These results demonstrate that [111In]PD-L1-mAb and NIR-PD-L1-mAb can detect graded levels of PD-L1 expression in human tumor xenografts in vivo. As a humanized antibody, these findings suggest clinical translation of radiolabeled versions of MPDL3280A for imaging. Specificity of NIR-PD-L1-mAb indicates the potential for optical imaging of PD-L1 expression in tumors in relevant pre-clinical as well as clinical settings. PMID:26848870

  19. Targeted positron emission tomography imaging of CXCR4 expression in patients with acute myeloid leukemia.

    PubMed

    Herhaus, Peter; Habringer, Stefan; Philipp-Abbrederis, Kathrin; Vag, Tibor; Gerngross, Carlos; Schottelius, Margret; Slotta-Huspenina, Julia; Steiger, Katja; Altmann, Torben; Weißer, Tanja; Steidle, Sabine; Schick, Markus; Jacobs, Laura; Slawska, Jolanta; Müller-Thomas, Catharina; Verbeek, Mareike; Subklewe, Marion; Peschel, Christian; Wester, Hans-Jürgen; Schwaiger, Markus; Götze, Katharina; Keller, Ulrich

    2016-08-01

    Acute myeloid leukemia originates from leukemia-initiating cells that reside in the protective bone marrow niche. CXCR4/CXCL12 interaction is crucially involved in recruitment and retention of leukemia-initiating cells within this niche. Various drugs targeting this pathway have entered clinical trials. To evaluate CXCR4 imaging in acute myeloid leukemia, we first tested CXCR4 expression in patient-derived primary blasts. Flow cytometry revealed that high blast counts in patients with acute myeloid leukemia correlate with high CXCR4 expression. The wide range of CXCR4 surface expression in patients was reflected in cell lines of acute myeloid leukemia. Next, we evaluated the CXCR4-specific peptide Pentixafor by positron emission tomography imaging in mice harboring CXCR4 positive and CXCR4 negative leukemia xenografts, and in 10 patients with active disease. [(68)Ga]Pentixafor-positron emission tomography showed specific measurable disease in murine CXCR4 positive xenografts, but not when CXCR4 was knocked out with CRISPR/Cas9 gene editing. Five of 10 patients showed tracer uptake correlating well with leukemia infiltration assessed by magnetic resonance imaging. The mean maximal standard uptake value was significantly higher in visually CXCR4 positive patients compared to CXCR4 negative patients. In summary, in vivo molecular CXCR4 imaging by means of positron emission tomography is feasible in acute myeloid leukemia. These data provide a framework for future diagnostic and theranostic approaches targeting the CXCR4/CXCL12-defined leukemia-initiating cell niche. PMID:27175029

  20. Targeted positron emission tomography imaging of CXCR4 expression in patients with acute myeloid leukemia

    PubMed Central

    Herhaus, Peter; Habringer, Stefan; Philipp-Abbrederis, Kathrin; Vag, Tibor; Gerngross, Carlos; Schottelius, Margret; Slotta-Huspenina, Julia; Steiger, Katja; Altmann, Torben; Weißer, Tanja; Steidle, Sabine; Schick, Markus; Jacobs, Laura; Slawska, Jolanta; Müller-Thomas, Catharina; Verbeek, Mareike; Subklewe, Marion; Peschel, Christian; Wester, Hans-Jürgen; Schwaiger, Markus; Götze, Katharina; Keller, Ulrich

    2016-01-01

    Acute myeloid leukemia originates from leukemia-initiating cells that reside in the protective bone marrow niche. CXCR4/CXCL12 interaction is crucially involved in recruitment and retention of leukemia-initiating cells within this niche. Various drugs targeting this pathway have entered clinical trials. To evaluate CXCR4 imaging in acute myeloid leukemia, we first tested CXCR4 expression in patient-derived primary blasts. Flow cytometry revealed that high blast counts in patients with acute myeloid leukemia correlate with high CXCR4 expression. The wide range of CXCR4 surface expression in patients was reflected in cell lines of acute myeloid leukemia. Next, we evaluated the CXCR4-specific peptide Pentixafor by positron emission tomography imaging in mice harboring CXCR4 positive and CXCR4 negative leukemia xenografts, and in 10 patients with active disease. [68Ga]Pentixafor-positron emission tomography showed specific measurable disease in murine CXCR4 positive xenografts, but not when CXCR4 was knocked out with CRISPR/Cas9 gene editing. Five of 10 patients showed tracer uptake correlating well with leukemia infiltration assessed by magnetic resonance imaging. The mean maximal standard uptake value was significantly higher in visually CXCR4 positive patients compared to CXCR4 negative patients. In summary, in vivo molecular CXCR4 imaging by means of positron emission tomography is feasible in acute myeloid leukemia. These data provide a framework for future diagnostic and theranostic approaches targeting the CXCR4/CXCL12-defined leukemia-initiating cell niche. PMID:27175029

  1. Aging-associated inflammation promotes selection for adaptive oncogenic events in B cell progenitors

    PubMed Central

    Henry, Curtis J.; Casás-Selves, Matias; Kim, Jihye; Zaberezhnyy, Vadym; Aghili, Leila; Daniel, Ashley E.; Jimenez, Linda; Azam, Tania; McNamee, Eoin N.; Clambey, Eric T.; Klawitter, Jelena; Serkova, Natalie J.; Tan, Aik Choon; Dinarello, Charles A.; DeGregori, James

    2015-01-01

    The incidence of cancer is higher in the elderly; however, many of the underlying mechanisms for this association remain unexplored. Here, we have shown that B cell progenitors in old mice exhibit marked signaling, gene expression, and metabolic defects. Moreover, B cell progenitors that developed from hematopoietic stem cells (HSCs) transferred from young mice into aged animals exhibited similar fitness defects. We further demonstrated that ectopic expression of the oncogenes BCR-ABL, NRASV12, or Myc restored B cell progenitor fitness, leading to selection for oncogenically initiated cells and leukemogenesis specifically in the context of an aged hematopoietic system. Aging was associated with increased inflammation in the BM microenvironment, and induction of inflammation in young mice phenocopied aging-associated B lymphopoiesis. Conversely, a reduction of inflammation in aged mice via transgenic expression of α-1-antitrypsin or IL-37 preserved the function of B cell progenitors and prevented NRASV12-mediated oncogenesis. We conclude that chronic inflammatory microenvironments in old age lead to reductions in the fitness of B cell progenitor populations. This reduced progenitor pool fitness engenders selection for cells harboring oncogenic mutations, in part due to their ability to correct aging-associated functional defects. Thus, modulation of inflammation — a common feature of aging — has the potential to limit aging-associated oncogenesis. PMID:26551682

  2. Alpha B-crystallin expression in mouse NIH 3T3 fibroblasts: glucocorticoid responsiveness and involvement in thermal protection.

    PubMed Central

    Aoyama, A; Fröhli, E; Schäfer, R; Klemenz, R

    1993-01-01

    alpha B-crystallin, a major soluble protein of vertebrate eye lenses, is a small heat shock protein which transiently accumulates in response to heat shock and other kinds of stress in mouse NIH 3T3 fibroblasts. Ectopic expression of an alpha B-crystallin cDNA clone renders NIH 3T3 cells thermoresistant. alpha B-crystallin accumulates in response to the synthetic glucocorticoid hormone dexamethasone. Dexamethasone-treated NIH 3T3 cells become thermoresistant to the same extent as they accumulate alpha B-crystallin. A cell clone in which alpha B-crystallin is superinduced upon heat shock acquires augmented thermotolerance. Expression of the ras oncogene causes a rapid but transient accumulation of alpha B-crystallin within 1 day. Later, sustained ras oncogene expression suppresses the dexamethasone-mediated alpha B-crystallin accumulation. Thus, oncogenic transformation triggered by the ras oncogene interferes with hormone-mediated accumulation of alpha B-crystallin and concomitant acquisition of thermoresistance. Other known heat shock proteins do not accumulate in response to ectopic alpha B-crystallin expression or to dexamethasone treatment. These results indicate that alpha B-crystallin can protect NIH 3T3 fibroblasts from thermal shock. Images PMID:8441415

  3. Compact whole-body fluorescent imaging of nude mice bearing EGFP expressing tumor

    NASA Astrophysics Data System (ADS)

    Chen, Yanping; Xiong, Tao; Chu, Jun; Yu, Li; Zeng, Shaoqun; Luo, Qingming

    2005-01-01

    Issue of tumor has been a hotspot of current medicine. It is important for tumor research to detect tumors bearing in animal models easily, fast, repetitively and noninvasivly. Many researchers have paid their increasing interests on the detecting. Some contrast agents, such as green fluorescent protein (GFP) and Discosoma red fluorescent protein (Dsred) were applied to enhance image quality. Three main kinds of imaging scheme were adopted to visualize fluorescent protein expressing tumors in vivo. These schemes based on fluorescence stereo microscope, cooled charge-coupled-device (CCD) or camera as imaging set, and laser or mercury lamp as excitation light source. Fluorescence stereo microscope, laser and cooled CCD are expensive to many institutes. The authors set up an inexpensive compact whole-body fluorescent imaging tool, which consisted of a Kodak digital camera (model DC290), fluorescence filters(B and G2;HB Optical, Shenyang, Liaoning, P.R. China) and a mercury 50-W lamp power supply (U-LH50HG;Olympus Optical, Japan) as excitation light source. The EGFP was excited directly by mercury lamp with D455/70 nm band-pass filter and fluorescence was recorded by digital camera with 520nm long-pass filter. By this easy operation tool, the authors imaged, in real time, fluorescent tumors growing in live mice. The imaging system is external and noninvasive. For half a year our experiments suggested the imaging scheme was feasible. Whole-body fluorescence optical imaging for fluorescent expressing tumors in nude mouse is an ideal tool for antitumor, antimetastatic, and antiangiogenesis drug screening.

  4. Fluorescence imaging of angiogenesis in green fluorescent protein-expressing tumors

    NASA Astrophysics Data System (ADS)

    Yang, Meng; Baranov, Eugene; Jiang, Ping; Li, Xiao-Ming; Wang, Jin W.; Li, Lingna; Yagi, Shigeo; Moossa, A. R.; Hoffman, Robert M.

    2002-05-01

    The development of therapeutics for the control of tumor angiogenesis requires a simple, reliable in vivo assay for tumor-induced vascularization. For this purpose, we have adapted the orthotopic implantation model of angiogenesis by using human and rodent tumors genetically tagged with Aequorea victoria green fluorescent protein (GFP) for grafting into nude mice. Genetically-fluorescent tumors can be readily imaged in vivo. The non-luminous induced capillaries are clearly visible against the bright tumor fluorescence examined either intravitally or by whole-body luminance in real time. Fluorescence shadowing replaces the laborious histological techniques for determining blood vessel density. High-level GFP-expressing tumor cell lines made it possible to acquire the high-resolution real-time fluorescent optical images of angiogenesis in both primary tumors and their metastatic lesions in various human and rodent tumor models by means of a light-based imaging system. Intravital images of angiogenesis onset and development were acquired and quantified from a GFP- expressing orthotopically-growing human prostate tumor over a 19-day period. Whole-body optical imaging visualized vessel density increasing linearly over a 20-week period in orthotopically-growing, GFP-expressing human breast tumor MDA-MB-435. Vessels in an orthotopically-growing GFP- expressing Lewis lung carcinoma tumor were visualized through the chest wall via a reversible skin flap. These clinically-relevant angiogenesis mouse models can be used for real-time in vivo evaluation of agents inhibiting or promoting tumor angiogenesis in physiological micro- environments.

  5. Human bladder carcinoma cell lines as indicators of oncogenic change relevant to urothelial neoplastic progression.

    PubMed Central

    Rieger, K. M.; Little, A. F.; Swart, J. M.; Kastrinakis, W. V.; Fitzgerald, J. M.; Hess, D. T.; Libertino, J. A.; Summerhayes, I. C.

    1995-01-01

    Analysis of human tumour-derived cell lines has previously resulted in the identification of novel transformation-related elements and provided a useful tool for functional studies of different genes. To establish the utility of such cell lines as indicators of change relevant to urothelial cancer, we have characterised the expression of five genes (p53, MDM2, Rb, E-cadherin, APC) within a panel of human bladder carcinoma cell lines. Using single-strand conformation polymorphism (SSCP) and direct sequencing, p53 mutations were identified in 7/15 (47%) cell lines reflecting events reported in bladder tumours. Immunohistochemical analysis of p53 in cultured cells and in paraffin-embedded sections of xenografts from the cell line panel revealed discordant results. An absence of p53 nuclear staining was associated with an exon 5 mutation in EJ and with multiple p53 mutations found in J82. Two cell lines positive for p53 staining in the absence of detectable mutation displayed overexpression of MDM2 (PSI, HT1197) in Western blot analysis. Loss or aberrant Rb expression was recorded in 5/15 (TCCSUP, SCaBER, 5637, HT1376, J82) cell lines. Absence of E-cadherin was recorded in 5/15 cell lines (TCCSUP, EJ, KK47, UM-UC-3, J82) with loss of alpha-catenin in immunoprecipitated E-cadherin complexes of CUBIII. Western blot analysis of APC revealed a truncated protein in 1/15 (CUBIII) cell lines. The characterisation of oncogenic events within this panel of human bladder carcinoma cell lines establishes a representation of change observed in bladder tumours and better defines the genotypic background in these experimental human cell models of neoplastic progression. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7669581

  6. Micro-positron emission tomography imaging of rat brain metabolism during expression of contextual conditioning.

    PubMed

    Luyten, Laura; Casteels, Cindy; Vansteenwegen, Debora; van Kuyck, Kris; Koole, Michel; Van Laere, Koen; Nuttin, Bart

    2012-01-01

    Using (18)F-fluorodeoxyglucose microPET imaging, we investigated the neurocircuitry of contextual anxiety versus control in awake, conditioned rats (n = 7-10 per group). In addition, we imaged a group expressing cued fear. Simultaneous measurements of startle amplitude and freezing time were used to assess conditioning. To the best of our knowledge, no neuroimaging studies in conditioned rats have been conducted thus far, although visualizing and quantifying the metabolism of the intact brain in behaving animals is clearly of interest. In addition, more insight into the neurocircuitry involved in contextual anxiety may stimulate the development of new treatments for anxiety disorders. Our main finding was hypermetabolism in a cluster comprising the bed nucleus of the stria terminalis (BST) in rats expressing contextual anxiety compared with controls. Analysis of a subset of rats showing the best behavioral results (n = 5 per subgroup) confirmed this finding. We also observed hypermetabolism in the same cluster in rats expressing contextual anxiety compared with rats expressing cued fear. Our results provide novel evidence for a role of the BST in the expression of contextual anxiety. PMID:22219287

  7. A new engineering approach to reveal correlation of physiological change and spontaneous expression from video images

    NASA Astrophysics Data System (ADS)

    Yang, Fenglei; Hu, Sijung; Ma, Xiaoyun; Hassan, Harnani; Wei, Dongqing

    2015-03-01

    Spontaneous expression is associated with physiological states, i.e., heart rate, respiration, oxygen saturation (SpO2%), and heart rate variability (HRV). There have yet not sufficient efforts to explore correlation of physiological change and spontaneous expression. This study aims to study how spontaneous expression is associated with physiological changes with an approved protocol or through the videos provided from Denver Intensity of Spontaneous Facial Action Database. Not like a posed expression, motion artefact in spontaneous expression is one of evitable challenges to be overcome in the study. To obtain a physiological signs from a region of interest (ROI), a new engineering approach is being developed with an artefact-reduction method consolidated 3D active appearance model (AAM) based track, affine transformation based alignment with opto-physiological mode based imaging photoplethysmography. Also, a statistical association spaces is being used to interpret correlation of spontaneous expressions and physiological states including their probability densities by means of Gaussian Mixture Model. The present work is revealing a new avenue of study associations of spontaneous expressions and physiological states with its prospect of applications on physiological and psychological assessment.

  8. Oncogenic roles of carbonic anhydrase 8 in human osteosarcoma cells.

    PubMed

    Wang, Tze-Kai; Lin, Yu-Ming; Lo, Che-Min; Tang, Chih-Hsin; Teng, Chieh-Lin Jerry; Chao, Wei-Ting; Wu, Min Huan; Liu, Chin-San; Hsieh, Mingli

    2016-06-01

    Carbonic anhydrase 8 (CA8), a member of the carbonic anhydrase family, is one of the three isozymes that do not catalyze the reversible hydration of carbon dioxide due to the lack of one important histidine. In the present study, we observed increased expression of CA8 in more aggressive types of human osteosarcoma (OS) cells and found that CA8 expression is correlated with disease stages, such that more intense expression occurs in the disease late stage. We also demonstrated that overexpression of CA8 in human OS (HOS) cells significantly increased cell proliferation both in vitro and in vivo. Downregulated CA8 sensitized cells to apoptotic stress induced by staurosporine and cisplatin, suggesting a specific role of CA8 to protect cells from stresses. In addition, downregulation of CA8 in HOS cells reduced cell invasion and colony formation ability in soft agar and further decreased matrix metalloproteinase 9 and focal adhesion kinase expression, indicating that CA8 might facilitate cancer cell invasion via the activation of FAK-MMP9 signaling. Interestingly, HOS cells with CA8 knockdown showed a significant decrease in glycolytic activity and cell death under glucose withdrawal, further indicating that CA8 may be involved in regulating aerobic glycolysis and enhancing cell viability. Knockdown of CA8 significantly decreased phosphorylated Akt expression suggesting that the oncogenic role of CA8 may be mediated by the regulation of Akt activation through p-Akt induction. Importantly, the inhibition of glycolysis by 2-deoxyglucose sensitized CA8 HOS-CA8-myc cells to cisplatin treatment under low glucose condition, highlighting a new therapeutic option for OS cancer. PMID:26711783

  9. 40 CFR 798.3320 - Combined chronic toxicity/oncogenicity.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 32 2014-07-01 2014-07-01 false Combined chronic toxicity/oncogenicity. 798.3320 Section 798.3320 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) TOXIC SUBSTANCES CONTROL ACT (CONTINUED) HEALTH EFFECTS TESTING GUIDELINES Chronic Exposure § 798.3320 Combined chronic toxicity/oncogenicity....

  10. The prolactin receptor mediates HOXA1-stimulated oncogenicity in mammary carcinoma cells.

    PubMed

    Hou, Lin; Xu, Bing; Mohankumar, Kumarasamypet M; Goffin, Vincent; Perry, Jo K; Lobie, Peter E; Liu, Dong-Xu

    2012-12-01

    The HOX genes are a highly conserved subgroup of homeodomain-containing transcription factors that are crucial to normal development. Forced expression of HOXA1 results in oncogenic transformation of immortalized human mammary cells with aggressive tumour formation in vivo. Microarray analysis identified that the prolactin receptor (PRLR) was significantly upregulated by forced expression of HOXA1 in mammary carcinoma cells. To determine prolactin (PRL) involvement in HOXA1‑induced oncogenicity in mammary carcinoma cells (MCF-7), we examined the effect of human prolactin (hPRL)-initiated PRLR signal transduction on changes in cellular behaviour mediated by HOXA1. Forced expression of HOXA1 in MCF-7 cells increased PRLR mRNA and protein expression. Forced expression of HOXA1 also enhanced hPRL-stimulated phosphorylation of both STAT5A/B and p44/42 MAPK, and increased subsequent transcriptional activity of STAT5A and STAT5B, and Elk-1 and Sap1a, respectively. Moreover, forced expression of HOXA1 in MCF-7 cells enhanced the hPRL‑stimulated increase in total cell number as a consequence of enhanced cell proliferation and cell survival, and also enhanced hPRL-stimulated anchorage-independent growth in soft agar. Increased anchorage-independent growth was attenuated by the PRLR antagonist ∆1-9-G129R‑hPRL. In conclusion, we have demonstrated that HOXA1 increases expression of the cell surface receptor PRLR and enhances PRLR-mediated signal transduction. Thus, the PRLR is one mediator of HOXA1‑stimulated oncogenicity in mammary carcinoma cells. PMID:23064471

  11. Oncometabolite Tinkers with Genome Folding, Boosting Oncogene Expression.

    PubMed

    Ing-Simmons, Elizabeth; Merkenschlager, Matthias

    2016-03-01

    A recent article makes a compelling case for a new mechanism by which heterozygous mutations in isocitrate dehydrogenases (IDH1/2) - implicated in cancer - undermine gene regulation. 2-Hydroxyglutarate (2HG) produced by mutant IDH alters the binding of the chromosomal organizer protein CTCF, disrupting the spatial and regulatory organization of the genome. PMID:26856236

  12. Enhanced MAF Oncogene Expression and Breast Cancer Bone Metastasis

    PubMed Central

    Pavlovic, Milica; Arnal-Estapé, Anna; Rojo, Federico; Bellmunt, Anna; Tarragona, Maria; Guiu, Marc; Planet, Evarist; Garcia-Albéniz, Xabier; Morales, Mónica; Urosevic, Jelena; Gawrzak, Sylwia; Rovira, Ana; Prat, Aleix; Nonell, Lara; Lluch, Ana; Jean-Mairet, Joël; Coleman, Robert; Albanell, Joan

    2015-01-01

    Background: There are currently no biomarkers for early breast cancer patient populations at risk of bone metastasis. Identification of mediators of bone metastasis could be of clinical interest. Methods: A de novo unbiased screening approach based on selection of highly bone metastatic breast cancer cells in vivo was used to determine copy number aberrations (CNAs) associated with bone metastasis. The CNAs associated with bone metastasis were examined in independent primary breast cancer datasets with annotated clinical follow-up. The MAF gene encoded within the CNA associated with bone metastasis was subjected to gain and loss of function validation in breast cancer cells (MCF7, T47D, ZR-75, and 4T1), its downstream mechanism validated, and tested in clinical samples. A multivariable Cox cause-specific hazard model with competing events (death) was used to test the association between 16q23 or MAF and bone metastasis. All statistical tests were two-sided. Results: 16q23 gain CNA encoding the transcription factor MAF mediates breast cancer bone metastasis through the control of PTHrP. 16q23 gain (hazard ratio (HR) for bone metastasis = 14.5, 95% confidence interval (CI) = 6.4 to 32.9, P < .001) as well as MAF overexpression (HR for bone metastasis = 2.5, 95% CI = 1.7 to 3.8, P < .001) in primary breast tumors were specifically associated with risk of metastasis to bone but not to other organs. Conclusions: These results suggest that MAF is a mediator of breast cancer bone metastasis. 16q23 gain or MAF protein overexpression in tumors may help to select patients at risk of bone relapse. PMID:26376684

  13. The LMO2 oncogene regulates DNA replication in hematopoietic cells

    PubMed Central

    Sincennes, Marie-Claude; Humbert, Magali; Grondin, Benoît; Lisi, Véronique; Veiga, Diogo F. T.; Haman, André; Cazaux, Christophe; Mashtalir, Nazar; Affar, EL Bachir; Verreault, Alain; Hoang, Trang

    2016-01-01

    Oncogenic transcription factors are commonly activated in acute leukemias and subvert normal gene expression networks to reprogram hematopoietic progenitors into preleukemic stem cells, as exemplified by LIM-only 2 (LMO2) in T-cell acute lymphoblastic leukemia (T-ALL). Whether or not these oncoproteins interfere with other DNA-dependent processes is largely unexplored. Here, we show that LMO2 is recruited to DNA replication origins by interaction with three essential replication enzymes: DNA polymerase delta (POLD1), DNA primase (PRIM1), and minichromosome 6 (MCM6). Furthermore, tethering LMO2 to synthetic DNA sequences is sufficient to transform these sequences into origins of replication. We next addressed the importance of LMO2 in erythroid and thymocyte development, two lineages in which cell cycle and differentiation are tightly coordinated. Lowering LMO2 levels in erythroid progenitors delays G1-S progression and arrests erythropoietin-dependent cell growth while favoring terminal differentiation. Conversely, ectopic expression in thymocytes induces DNA replication and drives these cells into cell cycle, causing differentiation blockade. Our results define a novel role for LMO2 in directly promoting DNA synthesis and G1-S progression. PMID:26764384

  14. CXCR4 in breast cancer: oncogenic role and therapeutic targeting

    PubMed Central

    Xu, Chao; Zhao, Hong; Chen, Haitao; Yao, Qinghua

    2015-01-01

    Chemokines are 8–12 kDa peptides that function as chemoattractant cytokines and are involved in cell activation, differentiation, and trafficking. Chemokines bind to specific G-protein-coupled seven-span transmembrane receptors. Chemokines play a fundamental role in the regulation of a variety of cellular, physiological, and developmental processes. Their aberrant expression can lead to a variety of human diseases including cancer. C-X-C chemokine receptor type 4 (CXCR4), also known as fusin or CD184, is an alpha-chemokine receptor specific for stromal-derived-factor-1 (SDF-1 also called CXCL12). CXCR4 belongs to the superfamily of the seven transmembrane domain heterotrimeric G protein-coupled receptors and is functionally expressed on the cell surface of various types of cancer cells. CXCR4 also plays a role in the cell proliferation and migration of these cells. Recently, CXCR4 has been reported to play an important role in cell survival, proliferation, migration, as well as metastasis of several cancers including breast cancer. This review is mainly focused on the current knowledge of the oncogenic role and potential drugs that target CXCR4 in breast cancer. Additionally, CXCR4 proangiogenic molecular mechanisms will be reviewed. Strict biunivocal binding affinity and activation of CXCR4/CXCL12 complex make CXCR4 a unique molecular target for prevention and treatment of breast cancer. PMID:26356032

  15. Oncogenic activation of ERG: A predominant mechanism in prostate cancer.

    PubMed

    Sreenath, Taduru L; Dobi, Albert; Petrovics, Gyorgy; Srivastava, Shiv

    2011-01-01

    Prevalent gene fusions involving regulatory sequences of the androgen receptor (AR) regulated genes (primarily TMPRSS2) and protein coding sequences of nuclear transcription factors of the ETS gene family (predominantly ERG) result in unscheduled androgen dependent ERG expression in prostate cancer (CaP).Cumulative data from a large number of studies in the past six years accentuate ERG alterations in more than half of all CaP patients in Western countries. Studies underscore that ERG functions are involved in the biology of CaP. ERG expression in normal context is selective to endothelial cells, specific hematopoetic cells and pre-cartilage cells. Normal functions of ERG are highlighted in hematopoetic stem cells. Emerging data continues to unravel molecular and cellular mechanisms by which ERG may contribute to CaP. Herein, we focus on biological and clinical aspects of ERG oncogenic alterations, potential of ERG-based stratification of CaP and the possibilities of targeting the ERG network in developing new therapeutic strategies for the disease. PMID:22279422

  16. Oncogene Induced Cellular Senescence Elicits an Anti-Warburg Effect

    PubMed Central

    Li, Mingxi; Durbin, Kenneth R.; Sweet, Steve M. M.; Tipton, Jeremiah D.; Zheng, Yupeng; Kelleher, Neil L.

    2013-01-01

    Cellular senescence, an irreversible cell cycle arrest induced by a diversity of stimuli, has been considered as an innate tumor suppressing mechanism with implications and applications in cancer therapy. Using a targeted proteomics approach we show that fibroblasts induced into senescence by expression of oncogenic Ras exhibit a decrease of global acetylation on all core histones, consistent with formation of senescence-associated heterochromatic foci. We also detected clear increases in repressive markers (e.g., >50% elevation of H3K27me2/3) along with decreases in histone marks associated with increased transcriptional expression/elongation (e.g., H3K36me2/3). Despite the increases in repressive marks of chromatin, 179 loci (of 2206 total) were found to be upregulated by global quantitative proteomics. The changes in the cytosolic proteome indicated an upregulation of mitochondrial proteins and downregulation of proteins involved in glycolysis. These alterations in primary metabolism are opposite of the well-known Warburg effect observed in cancer cells. This study significantly improves our understanding of stress-induced senescence and provides a potential application for triggering it in anti-proliferative strategies that target the primary metabolism in cancer cells. PMID:23798001

  17. A Computational Drug Repositioning Approach for Targeting Oncogenic Transcription Factors

    PubMed Central

    Gayvert, Kaitlyn; Dardenne, Etienne; Cheung, Cynthia; Boland, Mary Regina; Lorberbaum, Tal; Wanjala, Jackline; Chen, Yu; Rubin, Mark; Tatonetti, Nicholas P.; Rickman, David; Elemento, Olivier

    2016-01-01

    Summary Mutations in transcription factors (TFs) genes are frequently observed in tumors, often leading to aberrant transcriptional activity. Unfortunately, TFs are often considered undruggable due to the absence of targetable enzymatic activity. To address this problem, we developed CRAFTT, a Computational drug-Repositioning Approach For Targeting Transcription factor activity. CRAFTT combines ChIP-seq with drug-induced expression profiling to identify small molecules that can specifically perturb TF activity. Application to ENCODE ChIP-seq datasets revealed known drug-TF interactions and a global drug-protein network analysis further supported these predictions. Application of CRAFTT to ERG, a pro-invasive, frequently over-expressed oncogenic TF predicted that dexamethasone would inhibit ERG activity. Indeed, dexamethasone significantly decreased cell invasion and migration in an ERG-dependent manner. Furthermore, analysis of Electronic Medical Record data indicates a protective role for dexamethasone against prostate cancer. Altogether, our method provides a broadly applicable strategy to identify drugs that specifically modulate TF activity. PMID:27264179

  18. Disclosing the CXCR4 Expression in Lymphoproliferative Diseases by Targeted Molecular Imaging

    PubMed Central

    Wester, Hans Jürgen; Keller, Ulrich; Schottelius, Margret; Beer, Ambros; Philipp-Abbrederis, Kathrin; Hoffmann, Frauke; Šimeček, Jakub; Gerngross, Carlos; Lassmann, Michael; Herrmann, Ken; Pellegata, Natalia; Rudelius, Martina; Kessler, Horst; Schwaiger, Markus

    2015-01-01

    Chemokine ligand-receptor interactions play a pivotal role in cell attraction and cellular trafficking, both in normal tissue homeostasis and in disease. In cancer, chemokine receptor-4 (CXCR4) expression is an adverse prognostic factor. Early clinical studies suggest that targeting CXCR4 with suitable high-affinity antagonists might be a novel means for therapy. In addition to the preclinical evaluation of [68Ga]Pentixafor in mice bearing human lymphoma xenografts as an exemplary CXCR4-expressing tumor entity, we report on the first clinical applications of [68Ga]Pentixafor-Positron Emission Tomography as a powerful method for CXCR4 imaging in cancer patients. [68Ga]Pentixafor binds with high affinity and selectivity to human CXCR4 and exhibits a favorable dosimetry. [68Ga]Pentixafor-PET provides images with excellent specificity and contrast. This non-invasive imaging technology for quantitative assessment of CXCR4 expression allows to further elucidate the role of CXCR4/CXCL12 ligand interaction in the pathogenesis and treatment of cancer, cardiovascular diseases and autoimmune and inflammatory disorders. PMID:25825601

  19. Translation-dependent mechanisms lead to PML upregulation and mediate oncogenic K-RAS-induced cellular senescence

    PubMed Central

    Scaglioni, Pier Paolo; Rabellino, Andrea; Yung, Thomas M; Bernardi, Rosa; Choi, Sooyeon; Konstantinidou, Georgia; Nardella, Caterina; Cheng, Ke; Pandolfi, Pier Paolo

    2012-01-01

    Expression of oncogenic K-RAS in primary cells elicits oncogene-induced cellular senescence (OIS), a form of growth arrest that potently opposes tumourigenesis. This effect has been largely attributed to transcriptional mechanisms that depend on the p53 tumour suppressor protein. The PML tumour suppressor was initially identified as a component of the PML-RARα oncoprotein of acute promyelocytic leukaemia (APL). PML, a critical OIS mediator, is upregulated by oncogenic K-RAS in vivo and in vitro. We demonstrate here that oncogenic K-RAS induces PML protein upregulation by activating the RAS/MEK1/mTOR/eIF4E pathway even in the absence of p53. Under these circumstances, PML mRNA is selectively associated to polysomes. Importantly, we find that the PML 5′ untranslated mRNA region plays a key role in mediating PML protein upregulation and that its presence is essential for an efficient OIS response. These findings demonstrate that upregulation of PML translation plays a central role in oncogenic K-RAS-induced OIS. Thus, selective translation initiation plays a critical role in tumour suppression with important therapeutic implications for the treatment of solid tumours and APL. PMID:22359342

  20. JNK1 determines the oncogenic or tumor-suppressive activity of the integrin-linked kinase in human rhabdomyosarcoma.

    PubMed

    Durbin, Adam D; Somers, Gino R; Forrester, Michael; Pienkowska, Malgorzata; Hannigan, Gregory E; Malkin, David

    2009-06-01

    Although most reports describe the protein kinase integrin-linked kinase (ILK) as a proto-oncogene, occasional studies detail opposing functions in the regulation of normal and transformed cell proliferation, differentiation, and apoptosis. Here, we demonstrated that ILK functions as an oncogene in the highly aggressive pediatric sarcoma alveolar rhabdomyosarcoma (ARMS) and as a tumor suppressor in the related embryonal rhabdomyosarcoma (ERMS). These opposing functions hinge on signaling through a noncanonical ILK target, JNK1, to the proto-oncogene c-Jun. RNAi-mediated depletion of ILK induced activation of JNK and its target, c-Jun, resulting in growth of ERMS cells, whereas in ARMS cells, it led to loss of JNK/c-Jun signaling and suppression of growth both in vitro and in vivo. Ectopic expression of the fusion gene characteristic of ARMS (paired box 3-forkhead homolog in rhabdomyosarcoma [PAX3-FKHR]) in ERMS cells was sufficient to convert them to an ARMS signaling phenotype and render ILK activity oncogenic. Furthermore, restoration of JNK1 in ARMS reestablished a tumor-suppressive function for ILK. These findings indicate what we believe to be a novel effector pathway regulated by ILK, provide a mechanism for interconversion of oncogenic and tumor-suppressor functions of a single regulatory protein based on the genetic background of the tumor cells, and suggest a rationale for tailored therapy of rhabdomyosarcoma based on the different activities of ILK. PMID:19478459

  1. Hedgehog Cholesterolysis: Specialized Gatekeeper to Oncogenic Signaling

    PubMed Central

    Callahan, Brian P.; Wang, Chunyu

    2015-01-01

    Discussions of therapeutic suppression of hedgehog (Hh) signaling almost exclusively focus on receptor antagonism; however, hedgehog’s biosynthesis represents a unique and potentially targetable aspect of this oncogenic signaling pathway. Here, we review a key biosynthetic step called cholesterolysis from the perspectives of structure/function and small molecule inhibition. Cholesterolysis, also called cholesteroylation, generates cholesterol-modified Hh ligand via autoprocessing of a hedgehog precursor protein. Post-translational modification by cholesterol appears to be restricted to proteins in the hedgehog family. The transformation is essential for Hh biological activity and upstream of signaling events. Despite its decisive role in generating ligand, cholesterolysis remains conspicuously unexplored as a therapeutic target. PMID:26473928

  2. Mutational patterns in oncogenes and tumour suppressors.

    PubMed

    Baeissa, Hanadi M; Benstead-Hume, Graeme; Richardson, Christopher J; Pearl, Frances M G

    2016-06-15

    All cancers depend upon mutations in critical genes, which confer a selective advantage to the tumour cell. Knowledge of these mutations is crucial to understanding the biology of cancer initiation and progression, and to the development of targeted therapeutic strategies. The key to understanding the contribution of a disease-associated mutation to the development and progression of cancer, comes from an understanding of the consequences of that mutation on the function of the affected protein, and the impact on the pathways in which that protein is involved. In this paper we examine the mutation patterns observed in oncogenes and tumour suppressors, and discuss different approaches that have been developed to identify driver mutations within cancers that contribute to the disease progress. We also discuss the MOKCa database where we have developed an automatic pipeline that structurally and functionally annotates all proteins from the human proteome that are mutated in cancer. PMID:27284061

  3. Targeting the production of oncogenic microRNAs with multimodal synthetic small molecules.

    PubMed

    Vo, Duc Duy; Staedel, Cathy; Zehnacker, Laura; Benhida, Rachid; Darfeuille, Fabien; Duca, Maria

    2014-03-21

    MicroRNAs (miRNAs) are a recently discovered category of small RNA molecules that regulate gene expression at the post-transcriptional level. Accumulating evidence indicates that miRNAs are aberrantly expressed in a variety of human cancers and revealed to be oncogenic and to play a pivotal role in initiation and progression of these pathologies. It is now clear that the inhibition of oncogenic miRNAs, defined as blocking their biosynthesis or their function, could find an application in the therapy of different types of cancer in which these miRNAs are implicated. Here we report the design, synthesis, and biological evaluation of new small-molecule RNA ligands targeting the production of oncogenic microRNAs. In this work we focused our attention on miR-372 and miR-373 that are implicated in the tumorigenesis of different types of cancer such as gastric cancer. These two oncogenic miRNAs are overexpressed in gastric cancer cells starting from their precursors pre-miR-372 and pre-miR-373, two stem-loop structured RNAs that lead to mature miRNAs after cleavage by the enzyme Dicer. The small molecules described herein consist of the conjugation of two RNA binding motives, i.e., the aminoglycoside neomycin and different natural and artificial nucleobases, in order to obtain RNA ligands with increased affinity and selectivity compared to that of parent compounds. After the synthesis of this new series of RNA ligands, we demonstrated that they are able to inhibit the production of the oncogenic miRNA-372 and -373 by binding their pre-miRNAs and inhibiting the processing by Dicer. Moreover, we proved that some of these compounds bear anti-proliferative activity toward gastric cancer cells and that this activity is likely linked to a decrease in the production of targeted miRNAs. To date, only few examples of small molecules targeting oncogenic miRNAs have been reported, and such inhibitors could be extremely useful for the development of new anticancer therapeutic

  4. PET imaging of glioblastoma multiforme EGFR expression for therapeutic decision guidance.

    PubMed

    Wehrenberg-Klee, Eric; Redjal, Navid; Leece, Alicia; Turker, N Selcan; Heidari, Pedram; Shah, Khalid; Mahmood, Umar

    2015-01-01

    After initial therapy and total resection of glioblastoma multiforme (GBM), 80-90% of recurrences occur at the surgical margins. Insufficient sensitivity and specificity of current imaging techniques based on non-specific vascular imaging agents lead to delay in diagnosis of residual and/or recurrent disease. A tumor-specific imaging agent for GBM may improve detection of small residual disease in the post-operative period, and improve ability to distinguish tumor recurrence from its imaging mimics that can delay diagnosis. To this end, we developed an EGFR-targeted PET probe and assessed its ability to image EGFR WT (U87) and EGFRvIII (Gli36vIII) expressing GBMs in both murine intra-cranial xenografts and in a surgical-resection model. The developed imaging probe, (64)Cu-DOTAcetuximab-F(ab´)2, binds with a Kd of 11.2 nM to EGFR expressing GBM. (64)Cu-DOTA-cetuximab-F(ab´)2 specifically localized to intra-cranial tumor with a significant difference in SUVmean between tumor and contralateral brain for both Gli36vIII and U87 tumors (P<0.01 for both comparisons), with mean TBR of 22.5±0.7 for Gli36vIII tumors and 28.9±2.1 for U87 tumors (TBR±SEM). Tracer uptake by tumor was significantly inhibited by pre-injection with cetuximab (P<0.01 for both), with SUVmean reduced by 68% and 58% for Gli36vIII and U87 tumors, respectively. Surgical resection model PET-CT imaging demonstrates residual tumor and low nonspecific uptake in the resection site. We conclude that (64)Cu-DOTA-cetuximab-F(ab´)2 binds specifically to intracranial EGFR WT and EGFRvIII expressing GBM, demonstrates excellent TBR, and specifically images small residual tumor in a surgical model, suggesting future clinical utility in identifying true tumor recurrence. PMID:26269775

  5. PET imaging of glioblastoma multiforme EGFR expression for therapeutic decision guidance

    PubMed Central

    Wehrenberg-Klee, Eric; Redjal, Navid; Leece, Alicia; Turker, N Selcan; Heidari, Pedram; Shah, Khalid; Mahmood, Umar

    2015-01-01

    After initial therapy and total resection of glioblastoma multiforme (GBM), 80-90% of recurrences occur at the surgical margins. Insufficient sensitivity and specificity of current imaging techniques based on non-specific vascular imaging agents lead to delay in diagnosis of residual and/or recurrent disease. A tumor-specific imaging agent for GBM may improve detection of small residual disease in the post-operative period, and improve ability to distinguish tumor recurrence from its imaging mimics that can delay diagnosis. To this end, we developed an EGFR-targeted PET probe and assessed its ability to image EGFR WT (U87) and EGFRvIII (Gli36vIII) expressing GBMs in both murine intra-cranial xenografts and in a surgical-resection model. The developed imaging probe, 64Cu-DOTAcetuximab-F(ab´)2, binds with a Kd of 11.2 nM to EGFR expressing GBM. 64Cu-DOTA-cetuximab-F(ab´)2 specifically localized to intra-cranial tumor with a significant difference in SUVmean between tumor and contralateral brain for both Gli36vIII and U87 tumors (P<0.01 for both comparisons), with mean TBR of 22.5±0.7 for Gli36vIII tumors and 28.9±2.1 for U87 tumors (TBR±SEM). Tracer uptake by tumor was significantly inhibited by pre-injection with cetuximab (P<0.01 for both), with SUVmean reduced by 68% and 58% for Gli36vIII and U87 tumors, respectively. Surgical resection model PET-CT imaging demonstrates residual tumor and low nonspecific uptake in the resection site. We conclude that 64Cu-DOTA-cetuximab-F(ab´)2 binds specifically to intracranial EGFR WT and EGFRvIII expressing GBM, demonstrates excellent TBR, and specifically images small residual tumor in a surgical model, suggesting future clinical utility in identifying true tumor recurrence. PMID:26269775

  6. Real-Time Imaging of Gene Delivery and Expression with DNA Nanoparticle Technologies

    NASA Astrophysics Data System (ADS)

    Sun, Wenchao; Ziady, Assem G.

    The construction of safe, efficient, and modifiable synthetic DNA nanoparticles is an emerging technology that has achieved important milestones of success in the past 5 years. Advances in chemical conjugation, purification, and controlled synthesis have allowed researchers to produce uniform and stable particles, whose physical characteristics can be well characterized and monitored. As a result of these improvements, DNA nanoparticles have now been cleared for clinical testing, and show good potential for human gene therapy. A very important recent development in the study of DNA nanoparticles is the use of small-animal imaging. Real-time imaging has become a valuable technique for tracking particle biodistribution and gene transfer efficacy. In this chapter, we discuss how bioluminescent, positron emission tomography, and magnetic resonance imaging can be used separately or in concert to study particle delivery, localization, and magnitude of gene expression in vivo.

  7. Transglutaminase 2 contributes to a TP53-induced autophagy program to prevent oncogenic transformation

    PubMed Central

    Yeo, Shi Yun; Itahana, Yoko; Guo, Alvin Kunyao; Han, Rachel; Iwamoto, Kozue; Nguyen, Hung Thanh; Bao, Yi; Kleiber, Kai; Wu, Ya Jun; Bay, Boon Huat; Voorhoeve, Mathijs; Itahana, Koji

    2016-01-01

    Genetic alterations which impair the function of the TP53 signaling pathway in TP53 wild-type human tumors remain elusive. To identify new components of this pathway, we performed a screen for genes whose loss-of-function debilitated TP53 signaling and enabled oncogenic transformation of human mammary epithelial cells. We identified transglutaminase 2 (TGM2) as a putative tumor suppressor in the TP53 pathway. TGM2 suppressed colony formation in soft agar and tumor formation in a xenograft mouse model. The depletion of growth supplements induced both TGM2 expression and autophagy in a TP53-dependent manner, and TGM2 promoted autophagic flux by enhancing autophagic protein degradation and autolysosome clearance. Reduced expression of both CDKN1A, which regulates the cell cycle downstream of TP53, and TGM2 synergized to promote oncogenic transformation. Our findings suggest that TGM2-mediated autophagy and CDKN1A-mediated cell cycle arrest are two important barriers in the TP53 pathway that prevent oncogenic transformation. DOI: http://dx.doi.org/10.7554/eLife.07101.001 PMID:26956429

  8. Individual and Complementary Effects of Human Papillomavirus Oncogenes on Epithelial Cell Proliferation and Differentiation.

    PubMed

    Bergner, Sven; Halec, Gordana; Schmitt, Markus; Aubin, François; Alonso, Angel; Auvinen, Eeva

    2016-01-01

    Previous studies on human papillomavirus (HPV) type 16 protein functions have established the oncogenic nature of three viral proteins: E5, E6 and E7. Here we have studied the functions of these proteins by functional deletion of the individual E5, E6 or E7, or both E6 and E7 oncogenes in the context of the whole viral genome. These mutants, or the intact wild-type genome, were expressed from the natural viral promoters along with differentiation of epithelial HaCaT cells in three-dimensional collagen raft cultures. High episomal viral copy numbers were obtained using a transfection-based loxp-HPV16-eGFP-N1 vector system. All epithelial equivalents carrying the different HPV type 16 genomes showed pronounced hyperplastic and dysplastic morphology. Particularly the E7 oncogene, with contribution of E6, was shown to enhance cell proliferation. Specifically, the crucial role of E7 in HPV-associated hyperproliferation was clearly manifested. Based on morphological characteristics, immunohistochemical staining for differentiation and proliferation markers, and low expression of E1^E4, we propose that our raft culture models produce cervical intraepithelial neoplasia (CIN)1 and CIN2-like tissue. Our experimental setting provides an alternative tool to study concerted functions of HPV proteins in the development of epithelial dysplasia. PMID:26636751

  9. In vivo molecular imaging of chemokine receptor CXCR4 expression in patients with advanced multiple myeloma

    PubMed Central

    Philipp-Abbrederis, Kathrin; Herrmann, Ken; Knop, Stefan; Schottelius, Margret; Eiber, Matthias; Lückerath, Katharina; Pietschmann, Elke; Habringer, Stefan; Gerngroß, Carlos; Franke, Katharina; Rudelius, Martina; Schirbel, Andreas; Lapa, Constantin; Schwamborn, Kristina; Steidle, Sabine; Hartmann, Elena; Rosenwald, Andreas; Kropf, Saskia; Beer, Ambros J; Peschel, Christian; Einsele, Hermann; Buck, Andreas K; Schwaiger, Markus; Götze, Katharina; Wester, Hans-Jürgen; Keller, Ulrich

    2015-01-01

    CXCR4 is a G-protein-coupled receptor that mediates recruitment of blood cells toward its ligand SDF-1. In cancer, high CXCR4 expression is frequently associated with tumor dissemination and poor prognosis. We evaluated the novel CXCR4 probe [68Ga]Pentixafor for in vivo mapping of CXCR4 expression density in mice xenografted with human CXCR4-positive MM cell lines and patients with advanced MM by means of positron emission tomography (PET). [68Ga]Pentixafor PET provided images with excellent specificity and contrast. In 10 of 14 patients with advanced MM [68Ga]Pentixafor PET/CT scans revealed MM manifestations, whereas only nine of 14 standard [18F]fluorodeoxyglucose PET/CT scans were rated visually positive. Assessment of blood counts and standard CD34+ flow cytometry did not reveal significant blood count changes associated with tracer application. Based on these highly encouraging data on clinical PET imaging of CXCR4 expression in a cohort of MM patients, we conclude that [68Ga]Pentixafor PET opens a broad field for clinical investigations on CXCR4 expression and for CXCR4-directed therapeutic approaches in MM and other diseases. PMID:25736399

  10. Different Roles of Negative and Positive Components of the Circadian Clock in Oncogene-induced Neoplastic Transformation.

    PubMed

    Katamune, Chiharu; Koyanagi, Satoru; Shiromizu, Shoya; Matsunaga, Naoya; Shimba, Shigeki; Shibata, Shigenobu; Ohdo, Shigehiro

    2016-05-13

    In mammals, circadian rhythms in physiological function are generated by a molecular oscillator driven by transcriptional-translational feedback loop consisting of negative and positive regulators. Disruption of this circadian clock machinery is thought to increase the risk of cancer development, but the potential contributions of each component of circadian clock to oncogenesis have been little explored. Here we reported that negative and positive transcriptional regulators of circadian feedback loop had different roles in oncogene-induced neoplastic transformation. Mouse embryonic fibroblasts prepared from animals deficient in negative circadian clock regulators, Period2 (Per2) or Cryptochrome1/2 (Cry1/2), were prone to transformation induced by co-expression of H-ras(V12) and SV40 large T antigen (SV40LT). In contrast, mouse embryonic fibroblasts prepared from mice deficient in positive circadian clock regulators, Bmal1 or Clock, showed resistance to oncogene-induced transformation. In Per2 mutant and Cry1/2-null cells, the introduction of oncogenes induced expression of ATF4, a potent repressor of cell senescence-associated proteins p16INK4a and p19ARF. Elevated levels of ATF4 were sufficient to suppress expression of these proteins and drive oncogenic transformation. Conversely, in Bmal1-null and Clock mutant cells, the expression of ATF4 was not induced by oncogene introduction, which allowed constitutive expression of p16INK4a and p19ARF triggering cellular senescence. Although genetic ablation of either negative or positive transcriptional regulators of the circadian clock leads to disrupted rhythms in physiological functions, our findings define their different contributions to neoplastic cellular transformation. PMID:26961881

  11. Mitochondrial Ca2+ Remodeling is a Prime Factor in Oncogenic Behavior

    PubMed Central

    Rimessi, Alessandro; Patergnani, Simone; Bonora, Massimo; Wieckowski, Mariusz R.; Pinton, Paolo

    2015-01-01

    Cancer is sustained by defects in the mechanisms underlying cell proliferation, mitochondrial metabolism, and cell death. Mitochondrial Ca2+ ions are central to all these processes, serving as signaling molecules with specific spatial localization, magnitude, and temporal characteristics. Mutations in mtDNA, aberrant expression and/or regulation of Ca2+-handling/transport proteins and abnormal Ca2+-dependent relationships among the cytosol, endoplasmic reticulum, and mitochondria can cause the deregulation of mitochondrial Ca2+-dependent pathways that are related to these processes, thus determining oncogenic behavior. In this review, we propose that mitochondrial Ca2+ remodeling plays a pivotal role in shaping the oncogenic signaling cascade, which is a required step for cancer formation and maintenance. We will describe recent studies that highlight the importance of mitochondria in inducing pivotal “cancer hallmarks” and discuss possible tools to manipulate mitochondrial Ca2+ to modulate cancer survival. PMID:26161362

  12. Role of the proto-oncogene Pokemon in cellular transformation and ARF repression.

    PubMed

    Maeda, Takahiro; Hobbs, Robin M; Merghoub, Taha; Guernah, Ilhem; Zelent, Arthur; Cordon-Cardo, Carlos; Teruya-Feldstein, Julie; Pandolfi, Pier Paolo

    2005-01-20

    Aberrant transcriptional repression through chromatin remodelling and histone deacetylation has been postulated to represent a driving force underlying tumorigenesis because histone deacetylase inhibitors have been found to be effective in cancer treatment. However, the molecular mechanisms by which transcriptional derepression would be linked to tumour suppression are poorly understood. Here we identify the transcriptional repressor Pokemon (encoded by the Zbtb7 gene) as a critical factor in oncogenesis. Mouse embryonic fibroblasts lacking Zbtb7 are completely refractory to oncogene-mediated cellular transformation. Conversely, Pokemon overexpression leads to overt oncogenic transformation both in vitro and in vivo in transgenic mice. Pokemon can specifically repress the transcription of the tumour suppressor gene ARF through direct binding. We find that Pokemon is aberrantly overexpressed in human cancers and that its expression levels predict biological behaviour and clinical outcome. Pokemon's critical role in cellular transformation makes it an attractive target for therapeutic intervention. PMID:15662416

  13. Artistic expression in the development of new technology for three dimensional imaging

    NASA Astrophysics Data System (ADS)

    Oliveira, Sandra; Richardson, Martin; Azevedo, Isabel

    2011-02-01

    The medium of holography offers many new creative possibilities for the development of kinetic art. In the search of new forms of image display we examine new methods of capturing Three Dimensional animated images. Expression in new forms of visualisation leads to new methods for exploration auto-stereoscopic displays, three-dimensional imaging and holography. Artists have long combined cameras together to explore the human body as form and space and this paper sets out the potential of some of these techniques and in particular the technical potential of the use of multicamera capture techniques. Artists such as Tim Macmillan1 (2010) and Dayton Taylor2 (1997) use multi-lens cameras to create the illusion of capturing space and time for different effects, such as frozen moment, live action and slow motion (often seen as a cinemagraphic effect). However, their results are two-dimensional images made with a two-dimensional image capture system. Previous research on the interaction between art and technology has been based on twodimensional video art. This paper outlines a method of three-dimensional video capture to explore three-dimensional space and the human body. The stereoscopic specialist Ray Zone3 has written about the evolution of 3-D technology and 3-D film. Zone examines the development of these 3-D techniques and demonstrates the connection between two fields. This research extends our knowledge of Three Dimensional moving image as an art form. In the new art world, Holography has become a method increasingly used to develop kinetic art. In the search of new forms of display and image capture, we examine new techniques such as 3-D, including auto-stereoscopic display, three-dimensional imaging and holography.

  14. RUNX3 Has an Oncogenic Role in Head and Neck Cancer

    PubMed Central

    Tsunematsu, Takaaki; Kudo, Yasusei; Iizuka, Shinji; Ogawa, Ikuko; Fujita, Tsuyoshi; Kurihara, Hidemi; Abiko, Yoshimitsu; Takata, Takashi

    2009-01-01

    Background Runt-related transcription factor 3 (RUNX3) is a tumor suppressor of cancer and appears to be an important component of the transforming growth factor-beta (TGF-ß)-induced tumor suppression pathway. Surprisingly, we found that RUNX3 expression level in head and neck squamous cell carcinoma (HNSCC) tissues, which is one of the most common types of human cancer, was higher than that in normal tissues by a previously published microarray dataset in our preliminary study. Therefore, here we examined the oncogenic role of RUNX3 in HNSCC. Principal Findings Frequent RUNX3 expression and its correlation with malignant behavior were observed in HNSCC. Ectopic RUNX3 overexpression promoted cell growth and inhibited serum starvation-induced apoptosis and chemotherapeutic drug induced apoptosis in HNSCC cells. These findings were confirmed by RUNX3 knockdown. Moreover, RUNX3 overexpression enhanced tumorsphere formation. RUNX3 expression level was well correlated with the methylation status in HNSCC cells. Moreover, RUNX3 expression was low due to the methylation of its promoter in normal oral epithelial cells. Conclusions/Significance Our findings suggest that i) RUNX3 has an oncogenic role in HNSCC, ii) RUNX3 expression observed in HNSCC may be caused in part by demethylation during cancer development, and iii) RUNX3 expression can be a useful marker for predicting malignant behavior and the effect of chemotherapeutic drugs in HNSCC. PMID:19521519

  15. MicroRNAs Involved in Tumor Suppressor and Oncogene Pathways; Implications for Hepatobiliary Neoplasia

    PubMed Central

    Mott, Justin L.

    2009-01-01

    MicroRNAs are a class of small regulatory RNAs that function to modulate protein expression. This control allows for fine-tuning of the cellular phenotype, including regulation of proliferation, cell signaling, and apoptosis; not surprisingly, microRNAs contribute to liver cancer biology. Recent investigations in human liver cancers and tumor-derived cell lines have demonstrated decreased or increased expression of particular microRNAs in hepatobiliary cancer cells. Based on predicted and validated protein targets as well as functional consequences of altered expression, microRNAs with decreased expression in liver tumor cells may normally aid in limiting neoplastic transformation. Conversely, selected microRNAs that are upregulated in liver tumor cells can promote malignant features, contributing to carcinogenesis. In addition, microRNAs themselves are subject to regulated expression, including regulation by tumor suppressor and oncogene pathways. This review will focus on the expression and function of cancer-related microRNAs, including their intimate involvement in tumor suppressor and oncogene signaling networks relevant to hepatobiliary neoplasia. PMID:19585622

  16. Hepatitis C Virus Core from Two Different Genotypes Has an Oncogenic Potential but Is Not Sufficient for Transforming Primary Rat Embryo Fibroblasts in Cooperation with the H-ras Oncogene

    PubMed Central

    Chang, Jun; Yang, Se-Hwan; Cho, Young-Gyu; Hwang, Soon Bong; Hahn, Young Shin; Sung, Young Chul

    1998-01-01

    Persistent infection with hepatitis C virus (HCV) is associated with the development of liver cirrhosis and hepatocellular carcinoma. To examine the oncogenic potential of the HCV core gene product, primary rat embryo fibroblasts (REFs) were transfected with the core gene in the presence or absence of the H-ras oncogene. In contrast to a previous report (R. B. Ray, L. M. Lagging, K. Meyer, and R. Ray, J. Virol. 70:4438–4443, 1996), HCV core proteins from two different genotypes (type 1a and type 1b) were not found to transform REFs to tumorigenic phenotype in cooperation with the H-ras oncogene, although the core protein was successfully expressed 20 days after transfection. In addition, REFs transfected with E1A- but not core-expressing plasmid showed the phenotype of immortalized cells when selected with G418. The biological activity was confirmed by observing the transcription activation from two viral promoters, Rous sarcoma virus long terminal repeat and simian virus 40 promoter, which are known to be activated by the core protein from HCV-1 isolate. In contrast to the result with primary cells, the Rat-1 cell line, stably expressing HCV core protein, exhibited focus formation, anchorage-independent growth, and tumor formation in nude mice. HCV core protein was able to induce the transformation of Rat-1 cells with various efficiencies depending on the expression level of the core protein. These results indicate that HCV core protein has an oncogenic potential to transform the Rat-1 cell line but is not sufficient to either immortalize primary REFs by itself or transform primary cells in conjunction with the H-ras oncogene. PMID:9525629

  17. Oncogenic Ras stimulates Eiger/TNF exocytosis to promote growth

    PubMed Central

    Chabu, Chiswili; Xu, Tian

    2014-01-01

    Oncogenic mutations in Ras deregulate cell death and proliferation to cause cancer in a significant number of patients. Although normal Ras signaling during development has been well elucidated in multiple organisms, it is less clear how oncogenic Ras exerts its effects. Furthermore, cancers with oncogenic Ras mutations are aggressive and generally resistant to targeted therapies or chemotherapy. We identified the exocytosis component Sec15 as a synthetic suppressor of oncogenic Ras in an in vivo Drosophila mosaic screen. We found that oncogenic Ras elevates exocytosis and promotes the export of the pro-apoptotic ligand Eiger (Drosophila TNF). This blocks tumor cell death and stimulates overgrowth by activating the JNK-JAK-STAT non-autonomous proliferation signal from the neighboring wild-type cells. Inhibition of Eiger/TNF exocytosis or interfering with the JNK-JAK-STAT non-autonomous proliferation signaling at various steps suppresses oncogenic Ras-mediated overgrowth. Our findings highlight important cell-intrinsic and cell-extrinsic roles of exocytosis during oncogenic growth and provide a new class of synthetic suppressors for targeted therapy approaches. PMID:25411211

  18. Activation of ras oncogenes preceding the onset of neoplasia

    SciTech Connect

    Kumar, R.; Barbacid, M. ); Sukumar, S. )

    1990-06-01

    The identification of ras oncogenes in human and animal cancers including precancerous lesions indicates that these genes participate in the early stages of neoplastic development. Yet, these observations do not define the timing of ras oncogene activation in the multistep process of carcinogenesis. To ascertain the timing of ras oncogene activation, an animal model system was devised that involves the induction of mammary carcinomas in rats exposed at birth to the carcinogen nitrosomethylurea. High-resolution restriction fragment length polymorphism analysis of polymerase chain reaction-amplified ras sequences revealed the presence of both H-ras and K-ras oncogenes in normal mammary glands 2 weeks after carcinogen treatment and at least 2 months before the onset of neoplasia. These ras oncogenes can remain latent within the mammary gland until exposure to estrogens, demonstrating that activation of ras oncogenes can precede the onset of neoplasia and suggesting that normal physiological proliferative processes such as estrogen-induced mammary gland development may lead to neoplasia if the targeted cells harbor latent ras oncogenes.

  19. Non-invasive PET Imaging of PARP1 Expression in Glioblastoma Models

    PubMed Central

    Carney, Brandon; Carlucci, Giuseppe; Salinas, Beatriz; Di Gialleonardo, Valentina; Kossatz, Susanne; Vansteene, Axel; Longo, Valerie A.; Bolaender, Alexander; Chiosis, Gabriela; Keshari, Kayvan R.; Weber, Wolfgang A.; Reiner, Thomas

    2015-01-01

    Purpose The current study presents [18F]PARPi as imaging agent for PARP1 expression. Procedures [18F]PARPi was generated by conjugating a 2H-phthalazin-1-one scaffold to 4-[18F]fluorobenzoic acid. Biochemical assays, optical in vivo competition, biodistribution analysis, positron emission tomography (PET)/X-ray computed tomography, and PET/ magnetic resonance imaging studies were performed in subcutaneous and orthotopic mouse models of glioblastoma. Results [18F]PARPi shows suitable pharmacokinetic properties for brain tumor imaging (IC50=2.8±1.1 nM; logPCHI=2.15±0.41; plasma-free fraction=63.9±12.6 %) and accumulates selectively in orthotopic brain tumor tissue. Tracer accumulation in subcutaneous brain tumors was 1.82±0.21 %ID/g, whereas in healthy brain, the uptake was only 0.04±0.01 %ID/g. Conclusions [18F]PARPi is a selective PARP1 imaging agent that can be used to visualize glioblastoma in xenograft and orthotopic mouse models with high precision and good signal/noise ratios. It offers new opportunities to non-invasively image tumor growth and monitor interventions. PMID:26493053

  20. Multimodal molecular imaging system for pathway-specific reporter gene expression.

    PubMed

    Rossi, Marco; Massai, Luisa; Diamanti, Daniela; Fiengo, Pasquale; De Rosa, Antonella; Magrini, Roberta; Magnoni, Letizia; Chellini, Sara; Coniglio, Silvia; Diodato, Enrica; Pilli, Elena; Caradonna, Nicola Pasquale; Sardone, Gianluca; Monti, Martina; Roggeri, Riccardo; Lionetti, Vincenzo; Recchia, Fabio; Tunici, Patrizia; Valensin, Silvia; Scali, Carla; Pollio, Giuseppe; Porcari, Valentina

    2016-04-30

    Preclinical imaging modalities represent an essential tool to develop a modern and translational biomedical research. To date, Optical Imaging (OI) and Magnetic Resonance Imaging (MRI) are used principally in separate studies for molecular imaging studies. We decided to combine OI and MRI together through the development of a lentiviral vector to monitor the Wnt pathway response to Lithium Chloride (LiCl) treatment. The construct was stably infected in glioblastoma cells and, after intracranial transplantation in mice, serial MRI and OI imaging sessions were performed to detect human ferritin heavy chain protein (hFTH) and firefly luciferase enzyme (FLuc) respectively. The system allowed also ex vivo analysis using a constitutive fluorescence protein expression. In mice, LiCl administration has shown significantly increment of luminescence signal and a lower signal of T2 values (P<0.05), recorded noninvasively with OI and a 7 Tesla MRI scanner. This study indicates that OI and MRI can be performed in a single in vivo experiment, providing an in vivo proof-of-concept for drug discovery projects in preclinical phase. PMID:26987608

  1. Combining volumetric edge display and multiview display for expression of natural 3D images

    NASA Astrophysics Data System (ADS)

    Yasui, Ryota; Matsuda, Isamu; Kakeya, Hideki

    2006-02-01

    In the present paper the authors present a novel stereoscopic display method combining volumetric edge display technology and multiview display technology to realize presentation of natural 3D images where the viewers do not suffer from contradiction between binocular convergence and focal accommodation of the eyes, which causes eyestrain and sickness. We adopt volumetric display method only for edge drawing, while we adopt stereoscopic approach for flat areas of the image. Since focal accommodation of our eyes is affected only by the edge part of the image, natural focal accommodation can be induced if the edges of the 3D image are drawn on the proper depth. The conventional stereo-matching technique can give us robust depth values of the pixels which constitute noticeable edges. Also occlusion and gloss of the objects can be roughly expressed with the proposed method since we use stereoscopic approach for the flat area. We can attain a system where many users can view natural 3D objects at the consistent position and posture at the same time in this system. A simple optometric experiment using a refractometer suggests that the proposed method can give us 3-D images without contradiction between binocular convergence and focal accommodation.

  2. Noncanonical Roles of the Immune System in Eliciting Oncogene Addiction

    PubMed Central

    Casey, Stephanie C.; Bellovin, David I.; Felsher, Dean W.

    2013-01-01

    Summary Cancer is highly complex. The magnitude of this complexity makes it highly surprising that even the brief suppression of an oncogene can sometimes result in rapid and sustained tumor regression illustrating that cancers can be “oncogene addicted” [1-10]. The essential implication is that oncogenes may not only fuel the initiation of tumorigenesis, but in some cases necessarily their surfeit of activation is paramaount to maintain a neoplastic state [11]. Oncogene suppression acutely restores normal physiological programs that effectively overrides secondary genetic events and a cancer collapses [12,13]. Oncogene addiction is mediated both through both tumor intrinsic cell-autonomous mechanisms including proliferative arrest, apoptosis, differentiation and cellular senescence [1,2,4,12] but also host-dependent mechanisms that interact with these tumor intrinsic programs [14,15]. Notably, oncogene inactivation elicits a host immune response that involves specific immune effectors and cytokines that facilitate a remodeling of the tumor microenvironment including the shut down of angiogenesis and the induction of cellular senescence of tumor cells [16]. Hence, immune effectors are critically involved in tumor initiation and prevention [17-19] and progression [20], but also appear to be essential to tumor regression upon oncogene inactivation [21-23]. The understanding how the inactivation of an oncogene elicits a systemic signal in the host that prompts a deconstruction of a tumor could have important implications. The combination of oncogene-targeted therapy together with immunomodulatory therapy may be ideal for the development of both a robust tumor intrinsic as well as immunological effectively leading to sustained tumor regression. PMID:23571026

  3. Therapeutic Evaluation of microRNAs by Molecular Imaging

    PubMed Central

    Sekar, Thillai V.; Mohanram, Ramkumar Kunga; Foygel, Kira; Paulmurugan, Ramasamy

    2013-01-01

    MicroRNAs (miRNAs) function as regulatory molecules of gene expression with multifaceted activities that exhibit direct or indirect oncogenic properties, which promote cell proliferation, differentiation, and the development of different types of cancers. Because of their extensive functional involvement in many cellular processes, under both normal and pathological conditions such as various cancers, this class of molecules holds particular interest for cancer research. MiRNAs possess the ability to act as tumor suppressors or oncogenes by regulating the expression of different apoptotic proteins, kinases, oncogenes, and other molecular mechanisms that can cause the onset of tumor development. In contrast to current cancer medicines, miRNA-based therapies function by subtle repression of gene expression on a large number of oncogenic factors, and therefore are anticipated to be highly efficacious. Given their unique mechanism of action, miRNAs are likely to yield a new class of targeted therapeutics for a variety of cancers. More than thousand miRNAs have been identified to date, and their molecular mechanisms and functions are well studied. Furthermore, they are established as compelling therapeutic targets in a variety of cellular complications. However, the notion of using them as therapeutic tool was proposed only recently, given that modern imaging methods are just beginning to be deployed for miRNA research. In this review, we present a summary of various molecular imaging methods, which are instrumental in revealing the therapeutic potential of miRNAs, especially in various cancers. Imaging methods have recently been developed for monitoring the expression levels of miRNAs and their target genes by fluorescence-, bioluminescence- and chemiluminescence-based imaging techniques. Mature miRNAs bind to the untranslated regions (UTRs) of the target mRNAs and regulate target genes expressions. This concept has been used for the development of fluorescent reporter

  4. The oncogenic action of ionizing radiation on rat skin

    SciTech Connect

    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.

  5. An animal model allowing controlled receptor expression for molecular ultrasound imaging.

    PubMed

    Saini, Reshu; Sorace, Anna G; Warram, Jason M; Mahoney, Marshall J; Zinn, Kurt R; Hoyt, Kenneth

    2013-01-01

    Reported in this study is an animal model system for evaluating targeted ultrasound (US) contrast agents binding using adenoviral (Ad) vectors to modulate cellular receptor expression. An Ad vector encoding an extracellular hemagglutinin (HA) epitope tag and a green fluorescent protein (GFP) reporter was used to regulate receptor expression. A low and high receptor density (in breast cancer tumor bearing mice) was achieved by varying the Ad dose with a low plaque forming unit (PFU) on day 1 and high PFU on day 2 of experimentation. Targeted US contrast agents, or microbubbles (MB), were created by conjugating either biotinylated anti-HA or IgG isotype control antibodies to the MB surface with biotin-streptavidin linkage. Targeted and control MBs were administered on both days of experimentation and contrast-enhanced US (CEUS) was performed on each mouse using MB flash destruction technique. Signal intensities from MBs retained within tumor vasculature were analyzed through a custom Matlab program. Results showed intratumoral enhancement attributable to targeted MB accumulation was significantly increased from the low Ad vector dosing and the high Ad vector dosing (p = 0.001). Control MBs showed no significant differences between day 1 and day 2 imaging (p = 0.96). Additionally, targeted MBs showed a 10.5-fold increase in intratumoral image intensity on day 1 and an 18.8-fold increase in image intensity on day 2 compared with their control MB counterparts. PMID:23122640

  6. Oncogenic KRAS and BRAF Drive Metabolic Reprogramming in Colorectal Cancer.

    PubMed

    Hutton, Josiah E; Wang, Xiaojing; Zimmerman, Lisa J; Slebos, Robbert J C; Trenary, Irina A; Young, Jamey D; Li, Ming; Liebler, Daniel C

    2016-09-01

    Metabolic reprogramming, in which altered utilization of glucose and glutamine supports rapid growth, is a hallmark of most cancers. Mutations in the oncogenes KRAS and BRAF drive metabolic reprogramming through enhanced glucose uptake, but the broader impact of these mutations on pathways of carbon metabolism is unknown. Global shotgun proteomic analysis of isogenic DLD-1 and RKO colon cancer cell lines expressing mutant and wild type KRAS or BRAF, respectively, failed to identify significant differences (at least 2-fold) in metabolic protein abundance. However, a multiplexed parallel reaction monitoring (PRM) strategy targeting 73 metabolic proteins identified significant protein abundance increases of 1.25-twofold in glycolysis, the nonoxidative pentose phosphate pathway, glutamine metabolism, and the phosphoserine biosynthetic pathway in cells with KRAS G13D mutations or BRAF V600E mutations. These alterations corresponded to mutant KRAS and BRAF-dependent increases in glucose uptake and lactate production. Metabolic reprogramming and glucose conversion to lactate in RKO cells were proportional to levels of BRAF V600E protein. In DLD-1 cells, these effects were independent of the ratio of KRAS G13D to KRAS wild type protein. A study of 8 KRAS wild type and 8 KRAS mutant human colon tumors confirmed the association of increased expression of glycolytic and glutamine metabolic proteins with KRAS mutant status. Metabolic reprogramming is driven largely by modest (<2-fold) alterations in protein expression, which are not readily detected by the global profiling methods most commonly employed in proteomic studies. The results indicate the superiority of more precise, multiplexed, pathway-targeted analyses to study functional proteome systems. Data are available through MassIVE Accession MSV000079486 at ftp://MSV000079486@massive.ucsd.edu. PMID:27340238

  7. Genome wide proteomics of ERBB2 and EGFR and other oncogenic pathways in inflammatory breast cancer.

    PubMed

    Zhang, Emma Yue; Cristofanilli, Massimo; Robertson, Fredika; Reuben, James M; Mu, Zhaomei; Beavis, Ronald C; Im, Hogune; Snyder, Michael; Hofree, Matan; Ideker, Trey; Omenn, Gilbert S; Fanayan, Susan; Jeong, Seul-Ki; Paik, Young-Ki; Zhang, Anna Fan; Wu, Shiaw-Lin; Hancock, William S

    2013-06-01

    In this study we selected three breast cancer cell lines (SKBR3, SUM149 and SUM190) with different oncogene expression levels involved in ERBB2 and EGFR signaling pathways as a model system for the evaluation of selective integration of subsets of transcriptomic and proteomic data. We assessed the oncogene status with reads per kilobase per million mapped reads (RPKM) values for ERBB2 (14.4, 400, and 300 for SUM149, SUM190, and SKBR3, respectively) and for EGFR (60.1, not detected, and 1.4 for the same 3 cell lines). We then used RNA-Seq data to identify those oncogenes with significant transcript levels in these cell lines (total 31) and interrogated the corresponding proteomics data sets for proteins with significant interaction values with these oncogenes. The number of observed interactors for each oncogene showed a significant range, e.g., 4.2% (JAK1) to 27.3% (MYC). The percentage is measured as a fraction of the total protein interactions in a given data set vs total interactors for that oncogene in STRING (Search Tool for the Retrieval of Interacting Genes/Proteins, version 9.0) and I2D (Interologous Interaction Database, version 1.95). This approach allowed us to focus on 4 main oncogenes, ERBB2, EGFR, MYC, and GRB2, for pathway analysis. We used bioinformatics sites GeneGo, PathwayCommons and NCI receptor signaling networks to identify pathways that contained the four main oncogenes and had good coverage in the transcriptomic and proteomic data sets as well as a significant number of oncogene interactors. The four pathways identified were ERBB signaling, EGFR1 signaling, integrin outside-in signaling, and validated targets of C-MYC transcriptional activation. The greater dynamic range of the RNA-Seq values allowed the use of transcript ratios to correlate observed protein values with the relative levels of the ERBB2 and EGFR transcripts in each of the four pathways. This provided us with potential proteomic signatures for the SUM149 and 190 cell lines

  8. Genome wide proteomics of ERBB2 and EGFR and other oncogenic pathways in inflammatory breast cancer

    PubMed Central

    Zhang, Emma Yue; Cristofanilli, Massimo; Robertson, Fredika; Reuben, James M; Mu, Zhaomei; Beavis, Ronald C.; Im, Hogune; Snyder, Michael; Hofree, Matan; Ideker, Trey; Omenn, Gilbert S.; Fanayan, Susan; Jeong, Seul-Ki; Paik, Young-ki; Zhang, Anna Fan; Wu, Shiaw-Lin; Hancock, William S.

    2014-01-01

    In this study we selected three breast cancer cell lines (SKBR3, SUM149 and SUM190) with different oncogene expression levels involved in ERBB2 and EGFR signaling pathways as a model system for the evaluation of selective integration of subsets of transcriptomic and proteomic data. We assessed the oncogene status with RPKM values (Reads Per Kilobase per Million mapped reads1) for ERBB2 (14.4, 400 and 300 for SUM149, SUM 190 and SKBR3 respectively and for EGFR 60.1, not detected and 1.4 for the same 3 cell lines. We then used RNA-Seq data to identify those oncogenes with significant transcript levels in these cell lines (total 31) and interrogated the corresponding proteomics data sets for proteins with significant interaction values with these oncogenes. The number of observed interactors for each oncogene showed a significant range, e.g. 4.2% (JAK1) to 27.3% (MYC). The percentage is measured as a fraction of the total protein interactions in a given data set vs. total interactors for that oncogene in STRING (Search Tool for the Retrieval of Interacting Genes/Proteins, version 9.0) and I2D (Interologous Interaction Database, version 1.95). This approach allowed us to focus on 4 main oncogenes, ERBB2, EGFR, MYC, and GRB2, for pathway analysis. We used the following bioinformatics sites, GeneGo, PathwayCommons and NCI receptor signaling networks to identify pathways which contained the four main oncogenes, had good coverage in the transcriptomic and proteomic data sets as well as significant number of oncogene interactors. The four pathways identified were ERBB signaling, EGFR1 signaling, integrin outside-in signaling, and validated targets of C-MYC transcriptional activation. The greater dynamic range of the RNA-Seq values allowed the use of transcript ratios to correlate observed protein values with the relative levels of the ERBB2 and EGFR transcripts in each of the four pathways. This provided us with potential proteomic signatures for the SUM149 and 190 cell

  9. Overexpressed homeobox B9 regulates oncogenic activities by transforming growth factor-β1 in gliomas

    SciTech Connect

    Fang, Liping; Xu, Yinghui; Zou, Lijuan

    2014-03-28

    Highlights: • HOXB9 is overexpressed in gliomas. • HOXB9 over expression had shorter survival time than down expression in gliomas. • HOXB9 stimulated the proliferation, migration and sphere formation of glioma cells. • Activation of TGF-β1 contributed to HOXB9-induced oncogenic activities. - Abstract: Glioma is the leading cause of deaths related to tumors in the central nervous system. The mechanisms of gliomagenesis remain elusive to date. Homeobox B9 (HOXB9) has a crucial function in the regulation of gene expression and cell survival, but its functions in glioma formation and development have yet to be elucidated. This study showed that HOXB9 expression in glioma tissues was significantly higher than that in nontumor tissues. Higher HOXB9 expression was also significantly associated with advanced clinical stage in glioma patients. HOXB9 overexpression stimulated the proliferation, migration, and sphere formation of glioma cells, whereas HOXB9 knockdown elicited an opposite effect. HOXB9 overexpression also increased the tumorigenicity of glioma cells in vivo. Moreover, the activation of transforming growth factor-β1 contributed to HOXB9-induced oncogenic activities. HOXB9 could be used as a predictable biomarker to be detected in different pathological and histological subtypes in glioma for diagnosis or prognosis.

  10. Cervical keratinocytes containing stably replicating extrachromosomal HPV-16 are refractory to transformation by oncogenic H-Ras

    PubMed Central

    Berger, Kristi L.; Barriga, Felicia; Lace, Michael J.; Turek, Lubomir P.; Zamba, Gideon J.; Domann, Frederick E.; Lee, John H.; Klingelhutz, Aloysius J.

    2007-01-01

    Ras expression in human epithelial cells with integrated HPV genomes has been shown to cause tumorigenic transformation. The effects of Ras in cells representing early stage HPV-associated disease (i.e., when HPV is extrachromosomal and the oncogenes are under control of native promoters) have not been examined. Here, we used human cervical keratinocyte cell lines containing stably replicating extrachromosomal HPV-16 and present the novel finding that these cells resist transformation by oncogenic H-Ras. Ras expression consistently diminished anchorageindependent growth (AI), reduced E6 and E7 expression, and caused p53 induction in these cells. Conversely, AI was enhanced or maintained in Ras-transduced cervical cells that were immortalized with a 16E6/E7 retrovirus, and minimal effects on E6 and E7 expression were observed. Ras expression with either episomal HPV-16 or LXSN-E6/E7 was insufficient for tumorigenic growth suggesting that other events are needed for tumorigenic transformation. In conclusion, our results indicate that Ras-mediated transformation depends on the context of HPV oncogene expression and that this is an important point to address when developing HPV tumor models. PMID:16945398

  11. Lysyl oxidase activity regulates oncogenic stress response and tumorigenesis.

    PubMed

    Wiel, C; Augert, A; Vincent, D F; Gitenay, D; Vindrieux, D; Le Calvé, B; Arfi, V; Lallet-Daher, H; Reynaud, C; Treilleux, I; Bartholin, L; Lelievre, E; Bernard, D

    2013-01-01

    Cellular senescence, a stable proliferation arrest, is induced in response to various stresses. Oncogenic stress-induced senescence (OIS) results in blocked proliferation and constitutes a fail-safe program counteracting tumorigenesis. The events that enable a tumor in a benign senescent state to escape from OIS and become malignant are largely unknown. We show that lysyl oxidase activity contributes to the decision to maintain senescence. Indeed, in human epithelial cell the constitutive expression of the LOX or LOXL2 protein favored OIS escape, whereas inhibition of lysyl oxidase activity was found to stabilize OIS. The relevance of these in vitro observations is supported by in vivo findings: in a transgenic mouse model of aggressive pancreatic ductal adenocarcinoma (PDAC), increasing lysyl oxidase activity accelerates senescence escape, whereas inhibition of lysyl oxidase activity was found to stabilize senescence, delay tumorigenesis, and increase survival. Mechanistically, we show that lysyl oxidase activity favors the escape of senescence by regulating the focal-adhesion kinase. Altogether, our results demonstrate that lysyl oxidase activity participates in primary tumor growth by directly impacting the senescence stability. PMID:24113189

  12. Transformation with Oncogenic Ras and the Simian Virus 40 T Antigens Induces Caspase-Dependent Sensitivity to Fatty Acid Biosynthetic Inhibition

    PubMed Central

    Xu, Shihao; Spencer, Cody M.

    2015-01-01

    drive the transformation of normal cells to the cancerous state. These oncogenic alterations induce metabolic changes and dependencies that can be targeted to kill cancerous cells. Here, we find that the cellular transformation resulting from combined expression of the SV40 early region with an oncogenic Ras allele is sufficient to induce cellular susceptibility to fatty acid biosynthetic inhibition. Inhibition of fatty acid biosynthesis in these cells resulted in programmed cell death, which could be rescued by supplementing the medium with nonsaturated fatty acids. Similar results were observed with the expression of oncogenic Ras in nontransformed breast epithelial cells. Combined, our results suggest that specific oncogenic alleles induce metabolic dependencies that can be exploited to selectively kill cancerous cells. PMID:25855740

  13. Biological activities of v-myc and rearranged c-myc oncogenes in rat fibroblast cells in culture.

    PubMed

    Mougneau, E; Lemieux, L; Rassoulzadegan, M; Cuzin, F

    1984-09-01

    Two distinct forms of the myc oncogene were assayed for their ability to induce, in cultured rat fibroblast cells, the alterations of cellular growth controls observed upon transfer of the gene of polyoma virus encoding only the large T protein (plt). Both of these rearranged myc genes and the plt gene had been previously shown to cooperate with ras oncogenes for transformation of rat embryo fibroblasts (REF) and were thought to induce the same early step ("immortalization") of the tumoral transformation pathway. We now report that these two different oncogenes elicite the same response in the following biological assays: (i) reduction of the requirements in serum factors for growth in culture of cells of the established FR3T3 line; (ii) expression of transformed properties in low serum medium after transfer into FR3T3 cells expressing only the middle T protein of polyoma virus (MTT lines); (iii) conferring on REF cells the ability to grow as clonal colonies after seeding at low cell density; (iv) conferring on REF cells the ability to grow continuously in cell culture. These congruent phenotypes suggest that the activities of the large T and myc proteins result in the induction of the same molecular events. These results also provide simple biological assays and selective systems for oncogenes of the myc class. PMID:6091107

  14. A protein tagging system for signal amplification in gene expression and fluorescence imaging

    PubMed Central

    Tanenbaum, Marvin E.; Gilbert, Luke A.; Qi, Lei S.; Weissman, Jonathan S.; Vale, Ronald D.

    2014-01-01

    Summary Signals in many biological processes can be amplified by recruiting multiple copies of regulatory proteins to a site of action. Harnessing this principle, we have developed a novel protein scaffold, a repeating peptide array termed SunTag, which can recruit multiple copies of an antibody-fusion protein. We show that the SunTag can recruit up to 24 copies of GFP, thereby enabling long-term imaging of single protein molecules in living cells. We also use the SunTag to create a potent synthetic transcription factor by recruiting multiple copies of a transcriptional activation domain to a nuclease-deficient CRISPR/Cas9 protein and demonstrate strong activation of endogenous gene expression and re-engineered cell behavior with this system. Thus, the SunTag provides a versatile platform for multimerizing proteins on a target protein scaffold and is likely to have many applications in imaging and in controlling biological outputs. PMID:25307933

  15. Dense Image Matching for Mars Express HRSC Imagery Based on Precise Point Prediction Method

    NASA Astrophysics Data System (ADS)

    Geng, X.; Xu, Q.; Miao, J.; Hou, Y. F.; Xing, S.; Lan, C. Z.

    2016-06-01

    Currently, Mars Express HRSC imagery is an essential data source to derive high accuracy Mars topographic data. In view of the characteristics of Martian surface satellite imagery, a dense image matching scheme for HRSC imagery based on precise point prediction method is proposed. The image matching strategies of our method are elaborated in detail. Based on the proposed method, DEM and DOM of Martian surface are derived and compared with those published by ESA. The experiment results show that the root mean square error in planar direction is about three pixels, while the root mean square error in height direction is about one pixel. Moreover, the mean square error in plane direction show a certain systematic error and the reasons are analysed. Experiment results also demonstrate that the point prediction accuracy for corresponding points is up to 1-3 pixels.

  16. Modulation of junction tension by tumor suppressors and proto-oncogenes regulates cell-cell contacts.

    PubMed

    Bosveld, Floris; Guirao, Boris; Wang, Zhimin; Rivière, Mathieu; Bonnet, Isabelle; Graner, François; Bellaïche, Yohanns

    2016-02-15

    Tumor suppressors and proto-oncogenes play crucial roles in tissue proliferation. Furthermore, de-regulation of their functions is deleterious to tissue architecture and can result in the sorting of somatic rounded clones minimizing their contact with surrounding wild-type (wt) cells. Defects in the shape of somatic clones correlate with defects in proliferation, cell affinity, cell-cell adhesion, oriented cell division and cortical contractility. Combining genetics, live-imaging, laser ablation and computer simulations, we aim to analyze whether distinct or similar mechanisms can account for the common role of tumor suppressors and proto-oncogenes in cell-cell contact regulation. In Drosophila epithelia, the tumor suppressors Fat (Ft) and Dachsous (Ds) regulate cell proliferation, tissue morphogenesis, planar cell polarity and junction tension. By analyzing the evolution over time of ft mutant cells and clones, we show that ft clones reduce their cell-cell contacts with the surrounding wt tissue in the absence of concomitant cell divisions and over-proliferation. This contact reduction depends on opposed changes of junction tensions in the clone bulk and its boundary with neighboring wt tissue. More generally, either clone bulk or boundary junction tension is modulated by the activation of Yorkie, Myc and Ras, yielding similar contact reductions with wt cells. Together, our data highlight mechanical roles for proto-oncogene and tumor suppressor pathways in cell-cell interactions. PMID:26811379

  17. PIK3CA is implicated as an oncogene in ovarian cancer

    SciTech Connect

    Shayesteh, Laleh; Lu, Yiling; Kuo, Wen-Lin; Baldocchi, Russell; Godfrey, Tony; Collins, Colin; Pinkel, Daniel; Powell, Bethan; Mills,Gordon B.; Gray, Joe W.

    1998-03-25

    Ovarian cancer is the leading cause of death from gynecological malignancy and the fourth leading cause of cancer death among American women, yet little is known about its molecular aetiology. Studies using comparative genomic hybridization (CGH) have revealed several regions of recurrent, abnormal, DNA sequence copy number that may encode genes involved in the genesis or progression of the disease. One region at 3q26 found to be increased in copy number in approximately 40 percent of ovarian and other cancers contains PIK3CA, which encodes the p110 a catalytic subunit of phosphatidylinositol 3-kinase(PI3-kinase). The association between PIK3CA copy number and PI3-kinase activity makes PIK3CA a candidate oncogene because a broad range of cancer-related functions have been associated with PI3-kinase mediated signaling. These include proliferation, glucose transport and catabolism, cell adhesion, apoptosis, RAS signaling and oncogenic transformation. In addition, downstream effectors of PI3-kinase,AKT1 and AKT2, have been found to be amplified or activated in human tumors, including ovarian cancer. We show here that PIK3CA is frequently increased in copy number in ovarian cancers, that the increased copy number is associated with increased PIK3CA transcription, p110 a protein expression and PI3-kinase activity and that treatment with the PI3-kinase inhibitor LY294002 decreases proliferation and increases apoptosis. Our observations suggest PIK3CA is an oncogene that has an important role in ovarian cancer.

  18. Emerging Roles of Agrobacterial Plant-Transforming Oncogenes in Plant Defense Reactions

    NASA Astrophysics Data System (ADS)

    Bulgakov, Victor P.; Inyushkina, Yuliya V.; Gorpenchenko, Tatiana Y.; Koren, Olga G.; Shkryl, Yuri N.; Zhuravlev, Yuri N.

    2009-01-01

    For recent years, engineering plant metabolic pathways by using rol genes looks promising in several aspects. New directions of rol-gene studies are highlighted in this work underlying the unique regulatory properties of the genes. It is known that following agrobacterial infection, the Agrobacterium rhizogenes rolA, rolB and rolC genes are transferred to plant genome, causing tumor formation and hairy root disease. In this report, we show mat these oncogenes are also involved in regulation of plant defense reactions, including the production of secondary metabolites. Situations occur where the rol genes perform their own critical function to regulate secondary metabolism by bypassing upstream plant control mechanisms and directing defense reactions via a "short cut." The rolC gene expressed in transformed plant cells is efficient in establishing an enhanced resistance of host cells to salt and temperature stresses. The emerging complexity of the rol-gene triggered effects and the involvement of signals generated by these genes in basic processes of cell biology such as calcium and ROS signaling indicate that the plant oncogenes, like some animal protooncogenes, use sophisticated strategies to affect cell growth and differentiation. The data raise the intriguing possibility that some components of plant and animal oncogene signaling pathways share common features.

  19. Oncogenic Ras differentially regulates metabolism and anoikis in extracellular matrix-detached cells.

    PubMed

    Mason, J A; Davison-Versagli, C A; Leliaert, A K; Pape, D J; McCallister, C; Zuo, J; Durbin, S M; Buchheit, C L; Zhang, S; Schafer, Z T

    2016-08-01

    In order for cancer cells to survive during metastasis, they must overcome anoikis, a caspase-dependent cell death process triggered by extracellular matrix (ECM) detachment, and rectify detachment-induced metabolic defects that compromise cell survival. However, the precise signals used by cancer cells to facilitate their survival during metastasis remain poorly understood. We have discovered that oncogenic Ras facilitates the survival of ECM-detached cancer cells by using distinct effector pathways to regulate metabolism and block anoikis. Surprisingly, we find that while Ras-mediated phosphatidylinositol (3)-kinase signaling is critical for rectifying ECM-detachment-induced metabolic deficiencies, the critical downstream effector is serum and glucocorticoid-regulated kinase-1 (SGK-1) rather than Akt. Our data also indicate that oncogenic Ras blocks anoikis by diminishing expression of the phosphatase PHLPP1 (PH Domain and Leucine-Rich Repeat Protein Phosphatase 1), which promotes anoikis through the activation of p38 MAPK. Thus, our study represents a novel paradigm whereby oncogene-initiated signal transduction can promote the survival of ECM-detached cells through divergent downstream effectors. PMID:26915296

  20. Development of neutralizing monoclonal antibodies for oncogenic human papillomavirus types 31, 33, 45, 52, and 58.

    PubMed

    Brown, Martha J; Seitz, Hanna; Towne, Victoria; Müller, Martin; Finnefrock, Adam C

    2014-04-01

    Human papillomavirus (HPV) is the etiological agent for all cervical cancers, a significant number of other anogenital cancers, and a growing number of head and neck cancers. Two licensed vaccines offer protection against the most prevalent oncogenic types, 16 and 18, responsible for approximately 70% of cervical cancer cases worldwide and one of these also offers protection against types 6 and 11, responsible for 90% of genital warts. The vaccines are comprised of recombinantly expressed major capsid proteins that self-assemble into virus-like particles (VLPs) and prevent infection by eliciting neutralizing antibodies. Adding the other frequently identified oncogenic types 31, 33, 45, 52, and 58 to a vaccine would increase the coverage against HPV-induced cancers to approximately 90%. We describe the generation and characterization of panels of monoclonal antibodies to these five additional oncogenic HPV types, and the selection of antibody pairs that were high affinity and type specific and recognized conformation-dependent neutralizing epitopes. Such characteristics make these antibodies useful tools for monitoring the production and potency of a prototype vaccine as well as monitoring vaccine-induced immune responses in the clinic. PMID:24574536

  1. Overview on how oncogenic Kras promotes pancreatic carcinogenesis by inducing low intracellular ROS levels.

    PubMed

    Kong, Bo; Qia, Chengjia; Erkan, Mert; Kleeff, Jörg; Michalski, Christoph W

    2013-01-01

    Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease without clearly known disease causes. Recent epidemiological and animal studies suggest that the supplementation of dietary antioxidants (e.g., vitamins C and E) decreases cancer risk, implying that increased reactive oxygen species (ROS) may play a role in pancreatic carcinogenesis. However, oncogenic Kras mutations (e.g., Kras(G12D)), which are present in more than 90% of PDAC, have been proven to foster low intracellular ROS levels. Here, oncogenic Kras activates expression of a series of anti-oxidant genes via Nrf2 (nuclear factor, erythroid derived 2, like 2) and also mediates an unusual metabolic pathway of glutamine to generate NADPH. This can then be used as the reducing power for ROS detoxification, leading collectively to low ROS levels in pancreatic pre-neoplastic cells and in cancer cells. In adult stem cells and cancer stem cells, low ROS levels have been associated with the formation of a proliferation-permissive intracellular environment and with perseverance of self-renewal capacities. Therefore, it is conceivable that low intracellular ROS levels may contribute significantly to oncogenic Kras-mediated PDAC formation. PMID:24062691

  2. Oncogenic Potential of Hepatitis C Virus Proteins

    PubMed Central

    Banerjee, Arup; Ray, Ratna B.; Ray, Ranjit

    2010-01-01

    Chronic hepatitis C virus (HCV) infection is a major risk factor for liver disease progression, and may lead to cirrhosis and hepatocellular carcinoma (HCC). The HCV genome contains a single-stranded positive sense RNA with a cytoplasmic lifecycle. HCV proteins interact with many host-cell factors and are involved in a wide range of activities, including cell cycle regulation, transcriptional regulation, cell proliferation, apoptosis, lipid metabolism, and cell growth promotion. Increasing experimental evidences suggest that HCV contributes to HCC by modulating pathways that may promote malignant transformation of hepatocytes. At least four of the 10 HCV gene products, namely core, NS3, NS5A and NS5B play roles in several potentially oncogenic pathways. Induction of both endoplasmic reticulum (ER) stress and oxidative stress by HCV proteins may also contribute to hepatocyte growth promotion. The current review identifies important functions of the viral proteins connecting HCV infections and potential for development of HCC. However, most of the putative transforming potentials of the HCV proteins have been defined in artificial cellular systems, and need to be established relevant to infection and disease models. The new insight into the mechanisms for HCV mediated disease progression may offer novel therapeutic targets for one of the most devastating human malignancies in the world today. PMID:21994721

  3. Differential Expression of Chemokine Receptors and their Roles in Cancer Imaging

    PubMed Central

    Nimmagadda, Sridhar

    2012-01-01

    Chemokine/chemokine receptor interactions play diverse roles in cell migration and homeostasis. Emerging evidence suggests that cancer cells co-opt chemokine networks for survival, proliferation, immune evasion, and metastasis. Most of the chemokine receptors are reported to be involved in tumor progression. Given their extensive implication in cancer progression, several chemokine receptor/ligand axes are considered as potential therapeutic targets. This review provides a survey of chemokine receptor expression in cancer and evaluates the potential of chemokine receptor imaging as a tool for molecular characterization of cancer. PMID:22662317

  4. Magnetic Resonance Imaging of Tumors Colonized with Bacterial Ferritin-Expressing Escherichia coli

    PubMed Central

    Scadeng, Miriam; Geissinger, Ulrike; Haddad, Daniel; Basse-Lüsebrink, Thomas C.; Gbureck, Uwe; Jakob, Peter; Szalay, Aladar A.

    2011-01-01

    Background Recent studies have shown that human ferritin can be used as a reporter of gene expression for magnetic resonance imaging (MRI). Bacteria also encode three classes of ferritin-type molecules with iron accumulation properties. Methods and Findings Here, we investigated whether these bacterial ferritins can also be used as MRI reporter genes and which of the bacterial ferritins is the most suitable reporter. Bacterial ferritins were overexpressed in probiotic E. coli Nissle 1917. Cultures of these bacteria were analyzed and those generating highest MRI contrast were further investigated in tumor bearing mice. Among members of three classes of bacterial ferritin tested, bacterioferritin showed the most promise as a reporter gene. Although all three proteins accumulated similar amounts of iron when overexpressed individually, bacterioferritin showed the highest contrast change. By site-directed mutagenesis we also show that the heme iron, a unique part of the bacterioferritin molecule, is not critical for MRI contrast change. Tumor-specific induction of bacterioferritin-expression in colonized tumors resulted in contrast changes within the bacteria-colonized tumors. Conclusions Our data suggest that colonization and gene expression by live vectors expressing bacterioferritin can be monitored by MRI due to contrast changes. PMID:21984917

  5. Real time imaging of mRNA expression dynamics in live cells using protein complementation methods

    NASA Astrophysics Data System (ADS)

    Meller, Amit

    2009-03-01

    Traditional methods for mRNA quantification in cells, such as northern blots, quantitative PCR or microarrays assays, require cell lysis and therefore do not preserve its dynamics. These methods cannot be used to probe the spatio-temporal localization of mRNA in cells, which provide useful information for a wide range biomolecular process, including RNA metabolizim, expression kinetics and RNA interference. To probe mRNA dynamics in live prokaryotic and eukaryotic cells, we develop a method, which exploit the strong affinity of the eukaryotic initiation factor 4A (eIF4A) to specific RNA aptamers. Two parts of the eIF4A are fused to a split Green Fluorescence Protein (GFP), and are expressed in the cells at high abundance. However, only when the RNA apatmer is also present, the two protein parts complement and become fluorescent. Thus, the fluorescent background remains low, allowing us to directly image the expression of mRNA molecules in live e-coli cells from its early onset, over hours. We find that the expression kinetics can be classified in one out of at least three forms, which also display distinct spatial distributions. I will discuss the possible biological origin for these distributions and their time evolution.

  6. Imaging receptor for advanced glycation end product expression in mouse model of hind limb ischemia

    PubMed Central

    2013-01-01

    Background The purpose of this study is to image the effect of diabetes on expression of receptor for advanced glycation endproducts (RAGE) in limb ischemia in live animals. Methods Male wild-type C57BL/6 mice were either made diabetic or left as control. Two months later, diabetic and non-diabetic mice underwent left femoral artery ligation. The right leg served as lesion control. Five days later, mice were injected with 15.1 ± 4.4 MBq 99mTc-anti-RAGE F(ab’)2 and 4 to 5 h later (blood pool clearance) underwent SPECT/CT imaging. At the completion of imaging, mice were euthanized, hind limbs counted and sectioned, and scans reconstructed. Regions of interest were drawn on serial transverse sections comprising the hind limbs and activity in millicuries summed and divided by the injected dose (ID). Quantitative histology was performed for RAGE staining and angiogenesis. Results Uptake of 99mTc-anti-RAGE F(ab')2 as %ID × 10−3 was higher in the left (ischemic) limbs for the diabetic mice (n = 8) compared to non-diabetic mice (n = 8) (1.20 ± 0.44% vs. 0.49 ± 0.40%; P = 0.0007) and corresponded to less angiogenesis in the diabetic mice. Uptake was also higher in the right limbs of diabetic compared to non-diabetic animals (0.82 ± 0.33% vs. 0.40 ± 0.14%; P = 0.0004). Conclusions These data show the feasibility of imaging and quantifying the effect of diabetes on RAGE expression in limb ischemia. PMID:23663412

  7. Oncogene-dependent apoptosis is mediated by caspase-9

    PubMed Central

    Fearnhead, Howard O.; Rodriguez, Joe; Govek, Eve-Ellen; Guo, Wenjun; Kobayashi, Ryuji; Hannon, Greg; Lazebnik, Yuri A.

    1998-01-01

    Understanding how oncogenic transformation sensitizes cells to apoptosis may provide a strategy to kill tumor cells selectively. We previously developed a cell-free system that recapitulates oncogene dependent apoptosis as reflected by activation of caspases, the core of the apoptotic machinery. Here, we show that this activation requires a previously identified apoptosis-promoting complex consisting of caspase-9, APAF-1, and cytochrome c. As predicted by the in vitro system, preventing caspase-9 activation blocked drug-induced apoptosis in cells sensitized by E1A, an adenoviral oncogene. Oncogenes, such as E1A, appear to facilitate caspase-9 activation by several mechanisms, including the control of cytochrome c release from the mitochondria. PMID:9811857

  8. Oncogenes: The Passport for Viral Oncolysis Through PKR Inhibition

    PubMed Central

    Fernandes, Janaina

    2016-01-01

    The transforming properties of oncogenes are derived from gain-of-function mutations, shifting cell signaling from highly regulated homeostatic to an uncontrolled oncogenic state, with the contribution of the inactivating mutations in tumor suppressor genes P53 and RB, leading to tumor resistance to conventional and target-directed therapy. On the other hand, this scenario fulfills two requirements for oncolytic virus infection in tumor cells: inactivation of tumor suppressors and presence of oncoproteins, also the requirements to engage malignancy. Several of these oncogenes have a negative impact on the main interferon antiviral defense, the double-stranded RNA-activated protein kinase (PKR), which helps viruses to spontaneously target tumor cells instead of normal cells. This review is focused on the negative impact of overexpression of oncogenes on conventional and targeted therapy and their positive impact on viral oncolysis due to their ability to inhibit PKR-induced translation blockage, allowing virion release and cell death. PMID:27486347

  9. Oncogenes: The Passport for Viral Oncolysis Through PKR Inhibition.

    PubMed

    Fernandes, Janaina

    2016-01-01

    The transforming properties of oncogenes are derived from gain-of-function mutations, shifting cell signaling from highly regulated homeostatic to an uncontrolled oncogenic state, with the contribution of the inactivating mutations in tumor suppressor genes P53 and RB, leading to tumor resistance to conventional and target-directed therapy. On the other hand, this scenario fulfills two requirements for oncolytic virus infection in tumor cells: inactivation of tumor suppressors and presence of oncoproteins, also the requirements to engage malignancy. Several of these oncogenes have a negative impact on the main interferon antiviral defense, the double-stranded RNA-activated protein kinase (PKR), which helps viruses to spontaneously target tumor cells instead of normal cells. This review is focused on the negative impact of overexpression of oncogenes on conventional and targeted therapy and their positive impact on viral oncolysis due to their ability to inhibit PKR-induced translation blockage, allowing virion release and cell death. PMID:27486347

  10. Cooperation between the polyomavirus middle-T-antigen gene and the human c-myc oncogene in a rat thyroid epithelial differentiated cell line: model of in vitro progression.

    PubMed Central

    Berlingieri, M T; Portella, G; Grieco, M; Santoro, M; Fusco, A

    1988-01-01

    Two rat thyroid epithelial differentiated cell lines, PC Cl 3 and PC myc, were infected with the polyoma murine leukemia virus (PyMLV) carrying the Middle-T-antigen gene of polyomavirus. After infection, both cell lines acquired the typical markers of neoplastic transformation; however, the PC myc cells showed a greater malignant phenotype. Furthermore, the thyroid differentiated functions were completely suppressed in PC myc cells transformed by PyMLV, whereas they were, at least partially, retained in PC Cl 3 cells transformed by PyMLV, and in particular, thyroglobulin synthesis and secretion were not affected at all. Since no differences in the expression of the middle-T-antigen gene were observed in the two PyMLV-transformed cell lines, the different properties shown by these two infected cell lines must be ascribed to the expression of the c-myc oncogene. Images PMID:2838744

  11. Mate choice for more melanin as a mechanism to maintain a functional oncogene

    PubMed Central

    Fernandez, André A.; Morris, Molly R.

    2008-01-01

    The mechanisms by which cancer evolves and persists in natural systems have been difficult to ascertain. In the Xiphophorus melanoma model, a functional oncogene (Xiphophorus melanoma receptor kinase Xmrk) has been maintained for several million years despite being deleterious and in an extremely unstable genomic region. Melanomas in Xiphophorus spp. fishes (platyfishes and swordtails) have been investigated since the 1920s, and, yet, positive selection that could explain the maintenance of Xmrk has not been found. Here, we show that Xiphophorus cortezi females from two populations prefer males with the spotted caudal (Sc) melanin pattern, which is associated with the presence of the Xmrk oncogene and serves as the site of melanoma formation within this species. Moreover, X. cortezi females prefer males with an enhanced Sc to males with a reduced Sc pattern. RT-PCR analysis confirms tissue-specific Xmrk expression within the Sc pattern in X. cortezi. Because of the association of Xmrk with the Sc pigment pattern and the fact that melanoma formation augments this visual signal, sexual selection appears to be maintaining this oncogene because of a mating preference for Sc, as well as the exaggeration of this male trait. At the individual level, decreases in viability and fecundity because of Xmrk and subsequent melanoma formation may be mitigated via increases in mate acquisition. At the population level, maintenance of this oncogene appears to be under frequency dependent selection, as we detected female preference for males without Sc in a third population that had higher frequencies of Sc in females. PMID:18757731

  12. MUC1 alters oncogenic events and transcription in human breast cancer cells

    PubMed Central

    Hattrup, Christine L; Gendler, Sandra J

    2006-01-01

    Introduction MUC1 is an oncoprotein whose overexpression correlates with aggressiveness of tumors and poor survival of cancer patients. Many of the oncogenic effects of MUC1 are believed to occur through interaction of its cytoplasmic tail with signaling molecules. As expected for a protein with oncogenic functions, MUC1 is linked to regulation of proliferation, apoptosis, invasion, and transcription. Methods To clarify the role of MUC1 in cancer, we transfected two breast cancer cell lines (MDA-MB-468 and BT-20) with small interfering (si)RNA directed against MUC1 and analyzed transcriptional responses and oncogenic events (proliferation, apoptosis and invasion). Results Transcription of several genes was altered after transfection of MUC1 siRNA, including decreased MAP2K1 (MEK1), JUN, PDGFA, CDC25A, VEGF and ITGAV (integrin αv), and increased TNF, RAF1, and MMP2. Additional changes were seen at the protein level, such as increased expression of c-Myc, heightened phosphorylation of AKT, and decreased activation of MEK1/2 and ERK1/2. These were correlated with cellular events, as MUC1 siRNA in the MDA-MB-468 line decreased proliferation and invasion, and increased stress-induced apoptosis. Intriguingly, BT-20 cells displayed similar levels of apoptosis regardless of siRNA, and actually increased proliferation after MUC1 siRNA. Conclusion These results further the growing knowledge of the role of MUC1 in transcription, and suggest that the regulation of MUC1 in breast cancer may be more complex than previously appreciated. The differences between these two cell lines emphasize the importance of understanding the context of cell-specific signaling events when analyzing the oncogenic functions of MUC1, and caution against generalizing the results of individual cell lines without adequate confirmation in intact biological systems. PMID:16846534

  13. ERBB2 oncogenicity: ERBIN helps to perform the job

    PubMed Central

    Mei, Lin; Borg, Jean-Paul

    2015-01-01

    ERBB2 (v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 2) is an oncogenic tyrosine kinase receptor that is overexpressed in breast cancer. Antibodies and inhibitors targeting ERBB2 are currently available, although therapeutic failures remain frequent. We discuss here recent data showing that the scaffold protein ERBB2IP (ERBB2 interacting protein, best known as ERBIN) regulates ERBB2 stability and may represent a future therapeutic target. PMID:27308480

  14. Know thy neighbor: stromal cells can contribute oncogenic signals

    NASA Technical Reports Server (NTRS)

    Tlsty, T. D.; Hein, P. W.

    2001-01-01

    Although the stroma within carcinogenic lesions is known to be supportive and responsive to tumors, new data increasingly show that the stroma also has a more active, oncogenic role in tumorigenesis. Stromal cells and their products can transform adjacent tissues in the absence of pre-existing tumor cells by inciting phenotypic and genomic changes in the epithelial cells. The oncogenic action of distinctive stromal components has been demonstrated through a variety of approaches, which provide clues about the cellular pathways involved.

  15. ERβ decreases the invasiveness of triple-negative breast cancer cells by regulating mutant p53 oncogenic function

    PubMed Central

    Bado, Igor; Nikolos, Fotis; Rajapaksa, Gayani; Gustafsson, Jan-Åke; Thomas, Christoforos

    2016-01-01

    Most (80%) of the triple-negative breast cancers (TNBCs) express mutant p53 proteins that acquire oncogenic activities including promoting metastasis. We previously showed that wild-type ERβ (ERβ1) impedes epithelial to mesenchymal transition (EMT) and decreases the invasiveness of TNBC cells. In the present study we searched for signaling pathways that ERβ1 uses to inhibit EMT and invasion in TNBC cells. We show that ERβ1 binds to and opposes the transcriptional activity of mutant p53 at the promoters of genes that regulate metastasis. p63 that transcriptionally cooperates with mutant p53 also binds to ERβ1. Downregulation of p63 represses the epithelial phenotype of ERβ1-expressing cells and alters the expression of mutant p53 target genes. These results describe a novel mechanism through which ERβ1 can disturb oncogenic signals to inhibit aggressiveness in TNBCs. PMID:26871946

  16. Systemic delivery of siRNA by actively targeted polyion complex micelles for silencing the E6 and E7 human papillomavirus oncogenes.

    PubMed

    Nishida, Haruka; Matsumoto, Yoko; Kawana, Kei; Christie, R James; Naito, Mitsuru; Kim, Beob Soo; Toh, Kazuko; Min, Hyun Su; Yi, Yu; Matsumoto, Yu; Kim, Hyun Jin; Miyata, Kanjiro; Taguchi, Ayumi; Tomio, Kensuke; Yamashita, Aki; Inoue, Tomoko; Nakamura, Hiroe; Fujimoto, Asaha; Sato, Masakazu; Yoshida, Mitsuyo; Adachi, Katsuyuki; Arimoto, Takahide; Wada-Hiraike, Osamu; Oda, Katsutoshi; Nagamatsu, Takeshi; Nishiyama, Nobuhiro; Kataoka, Kazunori; Osuga, Yutaka; Fujii, Tomoyuki

    2016-06-10

    Human papillomavirus (HPV) E6 and E7 oncogenes are essential for the immortalization and maintenance of HPV-associated cancer and are ubiquitously expressed in cervical cancer lesions. Small interfering RNA (siRNA) coding for E6 and E7 oncogenes is a promising approach for precise treatment of cervical cancer, yet a delivery system is required for systemic delivery to solid tumors. Here, an actively targeted polyion complex (PIC) micelle was applied to deliver siRNAs coding for HPV E6/E7 to HPV cervical cancer cell tumors in immune-incompetent tumor-bearing mice. A cell viability assay revealed that both HPV type 16 and 18 E6/E7 siRNAs (si16E6/E7 and si18E6/E7, respectively) interfered with proliferation of cervical cancer cell lines in an HPV type-specific manner. A fluorescence imaging biodistribution analysis further revealed that fluorescence dye-labeled siRNA-loaded PIC micelles efficiently accumulated within the tumor mass after systemic administration. Ultimately, intravenous injection of si16E6/E7 and si18E6/E7-loaded PIC micelles was found to significantly suppress the growth of subcutaneous SiHa and HeLa tumors, respectively. The specific activity of siRNA treatment was confirmed by the observation that p53 protein expression was restored in the tumors excised from the mice treated with si16E6/E7- and si18E6/E7-loaded PIC micelles for SiHa and HeLa tumors, respectively. Therefore, the actively targeted PIC micelle incorporating HPV E6/E7-coding siRNAs demonstrated its therapeutic potential against HPV-associated cancer. PMID:26979870

  17. RECQL4 helicase has oncogenic potential in sporadic breast cancers.

    PubMed

    Arora, Arvind; Agarwal, Devika; Abdel-Fatah, Tarek Ma; Lu, Huiming; Croteau, Deborah L; Moseley, Paul; Aleskandarany, Mohammed A; Green, Andrew R; Ball, Graham; Rakha, Emad A; Chan, Stephen Yt; Ellis, Ian O; Wang, Lisa L; Zhao, Yongliang; Balajee, Adayabalam S; Bohr, Vilhelm A; Madhusudan, Srinivasan

    2016-03-01

    RECQL4 helicase is a molecular motor that unwinds DNA, a process essential during DNA replication and DNA repair. Germ-line mutations in RECQL4 cause type II Rothmund-Thomson syndrome (RTS), characterized by a premature ageing phenotype and cancer predisposition. RECQL4 is widely considered to be a tumour suppressor, although its role in human breast cancer is largely unknown. As the RECQL4 gene is localized to chromosome 8q24, a site frequently amplified in sporadic breast cancers, we hypothesized that it may play an oncogenic role in breast tumourigenesis. To address this, we analysed large cohorts for gene copy number changes (n = 1977), mRNA expression (n = 1977) and protein level (n = 1902). Breast cancer incidence was also explored in 58 patients with type II RTS. DNA replication dynamics and chemosensitivity was evaluated in RECQL4-depleted breast cancer cells in vitro. Amplification or gain in gene copy number (30.6%), high-level mRNA expression (51%) and high levels of protein (23%) significantly associated with aggressive tumour behaviour, including lymph node positivity, larger tumour size, HER2 overexpression, ER-negativity, triple-negative phenotypes and poor survival. RECQL4 depletion impaired the DNA replication rate and increased chemosensitivity in cultured breast cancer cells. Thus, although recognized as a 'safe guardian of the genome', our data provide compelling evidence that RECQL4 is tumour promoting in established breast cancers. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:26690729

  18. SUMOylation Confers Posttranslational Stability on NPM-ALK Oncogenic Protein.

    PubMed

    Vishwamitra, Deeksha; Curry, Choladda V; Shi, Ping; Alkan, Serhan; Amin, Hesham M

    2015-09-01

    Nucleophosmin-anaplastic lymphoma kinase-expressing (NPM-ALK+) T-cell lymphoma is an aggressive form of cancer that commonly affects children and adolescents. The expression of NPM-ALK chimeric oncogene results from the chromosomal translocation t(2;5)(p23;q35) that causes the fusion of the ALK and NPM genes. This translocation generates the NPM-ALK protein tyrosine kinase that forms the constitutively activated NPM-ALK/NPM-ALK homodimers. In addition, NPM-ALK is structurally associated with wild-type NPM to form NPM/NPM-ALK heterodimers, which can translocate to the nucleus. The mechanisms that sustain the stability of NPM-ALK are not fully understood. SUMOylation is a posttranslational modification that is characterized by the reversible conjugation of small ubiquitin-like modifiers (SUMOs) with target proteins. SUMO competes with ubiquitin for substrate binding and therefore, SUMOylation is believed to protect target proteins from proteasomal degradation. Moreover, SUMOylation contributes to the subcellular distribution of target proteins. Herein, we found that the SUMOylation pathway is deregulated in NPM-ALK+ T-cell lymphoma cell lines and primary lymphoma tumors from patients. We also identified Lys24 and Lys32 within the NPM domain as the sites where NPM-ALK conjugates with SUMO-1 and SUMO-3. Importantly, antagonizing SUMOylation by the SENP1 protease decreased the accumulation of NPM-ALK and suppressed lymphoma cell viability, proliferation, and anchorage-independent colony formation. One possible mechanism for the SENP1-mediated decrease in NPM-ALK levels was the increase in NPM-ALK association with ubiquitin, which facilitates its degradation. Our findings propose a model in which aberrancies in SUMOylation contribute to the pathogenesis of NPM-ALK+ T-cell lymphoma. Unraveling such pathogenic mechanisms may lead to devising novel strategies to eliminate this aggressive neoplasm. PMID:26476082

  19. Oncogene- and drug resistance-associated alternative exon usage in acute myeloid leukemia (AML).

    PubMed

    Mohamed, Aminetou Mint; Balsat, Marie; Thenoz, Morgan; Koering, Catherine; Payen-Gay, Lea; Cheok, Meyling; Mortada, Hussein; Auboeuf, Didier; Pinatel, Christiane; El-Hamri, Mohamed; Dumontet, Charles; Cros, Emeline; Flandrin-Gresta, Pascale; Nibourel, Olivier; Preudhomme, Claude; Michallet, Mauricette; Thomas, Xavier; Nicolini, Franck; Solly, Françoise; Guyotat, Denis; Campos, Lydia; Wattel, Eric; Mortreux, Franck

    2016-01-19

    In addition to spliceosome gene mutations, oncogene expression and drug resistance in AML might influence exon expression. We performed exon-array analysis and exon-specific PCR (ESPCR) to identify specific landscapes of exon expression that are associated with DEK and WT1 oncogene expression and the resistance of AML cells to AraC, doxorubicin or azacitidine. Data were obtained for these five conditions through exon-array analysis of 17 cell lines and 24 patient samples and were extended through qESPCR of samples from 152 additional AML cases. More than 70% of AEUs identified by exon-array were technically validated through ESPCR. In vitro, 1,130 to 5,868 exon events distinguished the 5 conditions from their respective controls while in vivo 6,560 and 9,378 events distinguished chemosensitive and chemoresistant AML, respectively, from normal bone marrow. Whatever the cause of this effect, 30 to 80% of mis-spliced mRNAs involved genes unmodified at the whole transcriptional level. These AEUs unmasked new functional pathways that are distinct from those generated by transcriptional deregulation. These results also identified new putative pathways that could help increase the understanding of the effects mediated by DEK or WT1, which may allow the targeting of these pathways to prevent resistance of AML cells to chemotherapeutic agents. PMID:26284582

  20. MTDH is an oncogene in multiple myeloma, which is suppressed by Bortezomib treatment

    PubMed Central

    Yang, Hongbao; Feng, Zhenqing; Yang, Ye

    2016-01-01

    Metadherin (MTDH) is identified as an oncogene in multiple cancers including breast cancer, bladder cancer and endometrial cancer. However, the function of MTDH in multiple myeloma (MM) is still unexplored. In this study, we disclose that MTDH is an oncogene in MM. This is characterized by an elevation mRNA level of MTDH and chromosomal gain of MTDH locus in MM cells compared to normal samples. Moreover, MTDH expression significantly increased in MMSET translocation (MS) subgroup, one of the high-risk subgroups in MM, and was significantly correlated with MM patients' poor outcomes in Total Therapy 2 (TT2) cohort. Further knockdown of MTDH expression by shRNA in MM cells induced cell apoptosis, inhibited MM cells growth in vitro and decreased xenograft tumor formation in vivo. Interestingly, opposite to TT2, MM patients with high-MTDH expression showed favorable survival outcomes in the TT3 cohort, while Bortezomib treatment was the major difference between TT2 and TT3 cohort. Furthermore we proved that Bortezomib suppressed pre- and post-transcription levels of MTDH expression of MM cells in vitro and in vivo. Finally, our studies demonstrated that MTDH is a transcriptional gene of MMSET/NFκB /MYC signaling in MM cells, which is inhibited by Bortezomib treatment. PMID:26683226

  1. Identification of MYC-Dependent Transcriptional Programs in Oncogene-Addicted Liver Tumors.

    PubMed

    Kress, Theresia R; Pellanda, Paola; Pellegrinet, Luca; Bianchi, Valerio; Nicoli, Paola; Doni, Mirko; Recordati, Camilla; Bianchi, Salvatore; Rotta, Luca; Capra, Thelma; Ravà, Micol; Verrecchia, Alessandro; Radaelli, Enrico; Littlewood, Trevor D; Evan, Gerard I; Amati, Bruno

    2016-06-15

    Tumors driven by activation of the transcription factor MYC generally show oncogene addiction. However, the gene expression programs that depend upon sustained MYC activity remain unknown. In this study, we employed a mouse model of liver carcinoma driven by a reversible tet-MYC transgene, combined with chromatin immunoprecipitation and gene expression profiling to identify MYC-dependent regulatory events. As previously reported, MYC-expressing mice exhibited hepatoblastoma- and hepatocellular carcinoma-like tumors, which regressed when MYC expression was suppressed. We further show that cellular transformation, and thus initiation of liver tumorigenesis, were impaired in mice harboring a MYC mutant unable to associate with the corepressor protein MIZ1 (ZBTB17). Notably, switching off the oncogene in advanced carcinomas revealed that MYC was required for the continuous activation and repression of distinct sets of genes, constituting no more than half of all genes deregulated during tumor progression and an even smaller subset of all MYC-bound genes. Altogether, our data provide the first detailed analysis of a MYC-dependent transcriptional program in a fully developed carcinoma and offer a guide to identifying the critical effectors contributing to MYC-driven tumor maintenance. Cancer Res; 76(12); 3463-72. ©2016 AACR. PMID:27197165

  2. Dual-Labeled Near-Infrared/99mTc Imaging Probes Using PAMAM-Coated Silica Nanoparticles for the Imaging of HER2-Expressing Cancer Cells

    PubMed Central

    Yamaguchi, Haruka; Tsuchimochi, Makoto; Hayama, Kazuhide; Kawase, Tomoyuki; Tsubokawa, Norio

    2016-01-01

    We sought to develop dual-modality imaging probes using functionalized silica nanoparticles to target human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer cells and achieve efficient target imaging of HER2-expressing tumors. Polyamidoamine-based functionalized silica nanoparticles (PCSNs) for multimodal imaging were synthesized with near-infrared (NIR) fluorescence (indocyanine green (ICG)) and technetium-99m (99mTc) radioactivity. Anti-HER2 antibodies were bound to the labeled PCSNs. These dual-imaging probes were tested to image HER2-overexpressing breast carcinoma cells. In vivo imaging was also examined in breast tumor xenograft models in mice. SK-BR3 (HER2 positive) cells were imaged with stronger NIR fluorescent signals than that in MDA-MB231 (HER2 negative) cells. The increased radioactivity of the SK-BR3 cells was also confirmed by phosphor imaging. NIR images showed strong fluorescent signals in the SK-BR3 tumor model compared to muscle tissues and the MDA-MB231 tumor model. Automatic well counting results showed increased radioactivity in the SK-BR3 xenograft tumors. We developed functionalized silica nanoparticles loaded with 99mTc and ICG for the targeting and imaging of HER2-expressing cells. The dual-imaging probes efficiently imaged HER2-overexpressing cells. Although further studies are needed to produce efficient isotope labeling, the results suggest that the multifunctional silica nanoparticles are a promising vehicle for imaging specific components of the cell membrane in a dual-modality manner. PMID:27399687

  3. Dual-Labeled Near-Infrared/(99m)Tc Imaging Probes Using PAMAM-Coated Silica Nanoparticles for the Imaging of HER2-Expressing Cancer Cells.

    PubMed

    Yamaguchi, Haruka; Tsuchimochi, Makoto; Hayama, Kazuhide; Kawase, Tomoyuki; Tsubokawa, Norio

    2016-01-01

    We sought to develop dual-modality imaging probes using functionalized silica nanoparticles to target human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer cells and achieve efficient target imaging of HER2-expressing tumors. Polyamidoamine-based functionalized silica nanoparticles (PCSNs) for multimodal imaging were synthesized with near-infrared (NIR) fluorescence (indocyanine green (ICG)) and technetium-99m ((99m)Tc) radioactivity. Anti-HER2 antibodies were bound to the labeled PCSNs. These dual-imaging probes were tested to image HER2-overexpressing breast carcinoma cells. In vivo imaging was also examined in breast tumor xenograft models in mice. SK-BR3 (HER2 positive) cells were imaged with stronger NIR fluorescent signals than that in MDA-MB231 (HER2 negative) cells. The increased radioactivity of the SK-BR3 cells was also confirmed by phosphor imaging. NIR images showed strong fluorescent signals in the SK-BR3 tumor model compared to muscle tissues and the MDA-MB231 tumor model. Automatic well counting results showed increased radioactivity in the SK-BR3 xenograft tumors. We developed functionalized silica nanoparticles loaded with (99m)Tc and ICG for the targeting and imaging of HER2-expressing cells. The dual-imaging probes efficiently imaged HER2-overexpressing cells. Although further studies are needed to produce efficient isotope labeling, the results suggest that the multifunctional silica nanoparticles are a promising vehicle for imaging specific components of the cell membrane in a dual-modality manner. PMID:27399687

  4. Structural and functional domains of the myb oncogene: requirements for nuclear transport, myeloid transformation, and colony formation.

    PubMed Central

    Ibanez, C E; Lipsick, J S

    1988-01-01

    The v-myb oncogene of avian myeloblastosis virus causes acute myelomonocytic leukemia in vivo and transforms only myeloid cells in vitro. Its product, p48v-myb, is a nuclear protein of unknown function. To determine structure-function relationships for this protein, we constructed a series of deletion mutants of v-myb, expressed them in retroviral vectors, and studied their biochemical and biological properties. We used these mutants to identify two separate domains of p48v-myb which had distinct roles in its accumulation in the cell nucleus. We showed that the viral sequences which normally encode both termini of p48v-myb were dispensible for transformation. In contrast, both copies of the highly conserved v-myb amino-terminal repeat were required for transformation. We also identified a carboxyl-terminal domain of p48v-myb which was required for the growth of v-myb-transformed myeloblasts in soft agar but not for morphological transformation. Images PMID:2835503

  5. Imaging neuronal responses in slice preparations of vomeronasal organ expressing a genetically encoded calcium sensor.

    PubMed

    Ma, Limei; Haga-Yamanaka, Sachiko; Yu, Qingfeng Elden; Qiu, Qiang; Kim, Sangseong; Yu, C Ron

    2011-01-01

    The vomeronasal organ (VNO) detects chemosensory signals that carry information about the social, sexual and reproductive status of the individuals within the same species. These intraspecies signals, the pheromones, as well as signals from some predators, activate the vomeronasal sensory neurons (VSNs) with high levels of specificity and sensitivity. At least three distinct families of G-protein coupled receptors, V1R, V2R and FPR, are expressed in VNO neurons to mediate the detection of the chemosensory cues. To understand how pheromone information is encoded by the VNO, it is critical to analyze the response profiles of individual VSNs to various stimuli and identify the specific receptors that mediate these responses. The neuroepithelia of VNO are enclosed in a pair of vomer bones. The semi-blind tubular structure of VNO has one open end (the vomeronasal duct) connecting to the nasal cavity. VSNs extend their dendrites to the lumen part of the VNO, where the pheromone cues are in contact with the receptors expressed at the dendritic knobs. The cell bodies of the VSNs form pseudo-stratified layers with V1R and V2R expressed in the apical and basal layers respectively. Several techniques have been utilized to monitor responses of VSNs to sensory stimuli. Among these techniques, acute slice preparation offers several advantages. First, compared to dissociated VSNs, slice preparations maintain the neurons in their native morphology and the dendrites of the cells stay relatively intact. Second, the cell bodies of the VSNs are easily accessible in coronal slice of the VNO to allow electrophysiology studies and imaging experiments as compared to whole epithelium and whole-mount preparations. Third, this method can be combined with molecular cloning techniques to allow receptor identification. Sensory stimulation elicits strong Ca2+ influx in VSNs that is indicative of receptor activation. We thus develop transgenic mice that express G-CaMP2 in the olfactory sensory

  6. Quantitative bioluminescence imaging of transgene expression in intact porcine antral follicles in vitro

    PubMed Central

    2014-01-01

    Background The porcine oocyte maturation in vivo occurs within the ovarian follicle and is regulated by the interactions between oocytes and surrounding follicular components, including theca, granulosa, and cumulus cells, and follicular fluid. Therefore, the antral follicle is an essential microenvironment for efficient oocyte maturation and its developmental competence. Quantitative bioluminescence imaging of firefly luciferase reporter genes in an intact antral follicle would allow investigation of changes in cellular and molecular events and in the context of the whole follicles. In this study, we investigate factors influencing bioluminescence measurements as a first step towards developing a new bioluminescence imaging system for intact antral follicles. Methods We analyzed the time course of bioluminescence emitted from transfected living intact follicles using a cationic lipid mediated gene transfer method with increasing doses (1-3 μg) of firefly luciferase reporter gene (pGL4). In addition, a standard luciferase assay was used to confirm the luciferase expression in granulosa cells in the transfected intact antral follicles. Finally, the dose effects of substrate, D-luciferin, were determined for optimal quantitative bioluminescence imaging of intact porcine antral follicles in vitro. Results The level of luciferase activity of follicles with 3 μg pGL4 was significantly (P < 0.05) greater than the 1 μg and 2 μg groups at 1 min after D-luciferin injection. The bioluminescence intensity of transfected follicles reached a peak at 1 min, and then it was significantly (P < 0.05) reduced within 2 min after injection of D-luciferin; with the level of bioluminescence emission remained constant from 2.5 to 10 min. The bioluminescence emission was maximal with 300 μg of D-luciferin. Conclusions The results of this study suggested that the investigation of factors influencing bioluminescence measurements is a critical step toward developing a

  7. IL-33 Facilitates Oncogene Induced Cholangiocarcinoma in Mice by an IL-6 Sensitive Mechanism

    PubMed Central

    Yamada, Daisaku; Rizvi, Sumera; Razumilava, Nataliya; Bronk, Steven F.; Davila, Jaime I.; Champion, Mia D.; Borad, Mitesh J.; Bezerra, Jorge A.; Chen, Xin; Gores, Gregory J.

    2015-01-01

    Cholangiocarcinoma (CCA) is a lethal hepatobiliary neoplasm originating from the biliary apparatus. In humans, CCA risk factors include hepatobiliary inflammation and fibrosis. The recently identified IL-1 family member, IL-33, has been shown to be a biliary mitogen which also promotes liver inflammation and fibrosis. Our aim was to generate a mouse model of CCA mimicking the human disease. Ectopic oncogene expression in the biliary tract was accomplished by the Sleeping Beauty transposon transfection system with transduction of constitutively active AKT (myr-AKT) and Yes-associated protein (YAP). Intrabiliary instillation of the transposon-transposase complex was coupled with lobar bile duct ligation in CL57BL/6 mice, followed by administration of IL-33 for three consecutive days. Tumors developed in 72% of the male mice receiving both oncogenes plus IL-33 by 10 weeks, but in only 20% of the male mice transduced with the oncogenes alone. Tumors expressed SOX9 and pancytokeratin (PanCK) [features of cholangiocarcinoma] but were negative for HepPar1 [a marker of hepatocellular carcinoma (HCC)]. RNA profiling revealed substantive overlap with human CCA specimens. Not only did IL-33 induce IL-6 expression by human cholangiocytes, but IL-33 likely facilitated tumor development in vivo by an IL-6 sensitive process, as tumor development was significantly attenuated in Il-6 -/- male animals. Furthermore, tumor formation occurred at a similar rate when IL-6 was substituted for IL-33 in this model. In conclusion, the transposase-mediated transduction of constitutively active AKT and YAP in the biliary epithelium coupled with lobar obstruction and IL-33 administration results in the development of CCA with morphological and biochemical features of the human disease. This model highlights the role of inflammatory cytokines in CCA oncogenesis. PMID:25580681

  8. MMP-13 In-Vivo Molecular Imaging Reveals Early Expression in Lung Adenocarcinoma

    PubMed Central

    Salaün, Mathieu; Peng, Jing; Hensley, Harvey H.; Roder, Navid; Flieder, Douglas B.; Houlle-Crépin, Solène; Abramovici-Roels, Olivia; Sabourin, Jean-Christophe; Thiberville, Luc; Clapper, Margie L.

    2015-01-01

    Introduction Several matrix metalloproteinases (MMPs) are overexpressed in lung cancer and may serve as potential targets for the development of bioactivable probes for molecular imaging. Objective To characterize and monitor the activity of MMPs during the progression of lung adenocarcinoma. Methods K-rasLSL-G12D mice were imaged serially during the development of adenocarcinomas using fluorescence molecular tomography (FMT) and a probe specific for MMP-2, -3, -9 and -13. Lung tumors were identified using FMT and MRI co-registration, and the probe concentration in each tumor was assessed at each time-point. The expression of Mmp2, -3, -9, -13 was quantified by qRT-PCR using RNA isolated from microdissected tumor cells. Immunohistochemical staining of overexpressed MMPs in animals was assessed on human lung tumors. Results In mice, 7 adenomas and 5 adenocarcinomas showed an increase in fluorescent signal on successive FMT scans, starting between weeks 4 and 8. qRT-PCR assays revealed significant overexpression of only Mmp-13 in mice lung tumors. In human tumors, a high MMP-13 immunostaining index was found in tumor cells from invasive lesions (24/27), but in none of the non-invasive (0/4) (p=0.001). Conclusion MMP-13 is detected in early pulmonary invasive adenocarcinomas and may be a potential target for molecular imaging of lung cancer. PMID:26193700

  9. (68)Ga-Pentixafor-PET/CT for Imaging of Chemokine Receptor 4 Expression in Glioblastoma.

    PubMed

    Lapa, Constantin; Lückerath, Katharina; Kleinlein, Irene; Monoranu, Camelia Maria; Linsenmann, Thomas; Kessler, Almuth F; Rudelius, Martina; Kropf, Saskia; Buck, Andreas K; Ernestus, Ralf-Ingo; Wester, Hans-Jürgen; Löhr, Mario; Herrmann, Ken

    2016-01-01

    Chemokine receptor-4 (CXCR4) has been reported to be overexpressed in glioblastoma (GBM) and to be associated with poor survival. This study investigated the feasibility of non-invasive CXCR4-directed imaging with positron emission tomography/computed tomography (PET/CT) using the radiolabelled chemokine receptor ligand (68)Ga-Pentixafor. 15 patients with clinical suspicion on primary or recurrent glioblastoma (13 primary, 2 recurrent tumors) underwent (68)Ga-Pentixafor-PET/CT for assessment of CXCR4 expression prior to surgery. O-(2-(18)F-fluoroethyl)-L-tyrosine ((18)F-FET) PET/CT images were available in 11/15 cases and were compared visually and semi-quantitatively (SUVmax, SUVmean). Tumor-to-background ratios (TBR) were calculated for both PET probes. (68)Ga-Pentixafor-PET/CT results were also compared to histological CXCR4 expression on neuronavigated surgical samples. (68)Ga-Pentixafor-PET/CT was visually positive in 13/15 cases with SUVmean and SUVmax of 3.0±1.5 and 3.9±2.0 respectively. Respective values for (18)F-FET were 4.4±2.0 (SUVmean) and 5.3±2.3 (SUVmax). TBR for SUVmean and SUVmax were higher for (68)Ga-Pentixafor than for (18)F-FET (SUVmean 154.0±90.7 vs. 4.1±1.3; SUVmax 70.3±44.0 and 3.8±1.2, p<0.01), respectively. Histological analysis confirmed CXCR4 expression in tumor areas with high (68)Ga-Pentixafor uptake; regions of the same tumor without apparent (68)Ga-Pentixafor uptake showed no or low receptor expression. In this pilot study, (68)Ga-Pentixafor retention has been observed in the vast majority of glioblastoma lesions and served as readout for non-invasive determination of CXCR4 expression. Given the paramount importance of the CXCR4/SDF-1 axis in tumor biology, (68)Ga-Pentixafor-PET/CT might prove a useful tool for sensitive, non-invasive in-vivo quantification of CXCR4 as well as selection of patients who might benefit from CXCR4-directed therapy. PMID:26909116

  10. 68Ga-Pentixafor-PET/CT for Imaging of Chemokine Receptor 4 Expression in Glioblastoma

    PubMed Central

    Lapa, Constantin; Lückerath, Katharina; Kleinlein, Irene; Monoranu, Camelia Maria; Linsenmann, Thomas; Kessler, Almuth F.; Rudelius, Martina; Kropf, Saskia; Buck, Andreas K.; Ernestus, Ralf-Ingo; Wester, Hans-Jürgen; Löhr, Mario; Herrmann, Ken

    2016-01-01

    Chemokine receptor-4 (CXCR4) has been reported to be overexpressed in glioblastoma (GBM) and to be associated with poor survival. This study investigated the feasibility of non-invasive CXCR4-directed imaging with positron emission tomography/computed tomography (PET/CT) using the radiolabelled chemokine receptor ligand 68Ga-Pentixafor. 15 patients with clinical suspicion on primary or recurrent glioblastoma (13 primary, 2 recurrent tumors) underwent 68Ga-Pentixafor-PET/CT for assessment of CXCR4 expression prior to surgery. O-(2-18F-fluoroethyl)-L-tyrosine (18F-FET) PET/CT images were available in 11/15 cases and were compared visually and semi-quantitatively (SUVmax, SUVmean). Tumor-to-background ratios (TBR) were calculated for both PET probes. 68Ga-Pentixafor-PET/CT results were also compared to histological CXCR4 expression on neuronavigated surgical samples. 68Ga-Pentixafor-PET/CT was visually positive in 13/15 cases with SUVmean and SUVmax of 3.0±1.5 and 3.9±2.0 respectively. Respective values for 18F-FET were 4.4±2.0 (SUVmean) and 5.3±2.3 (SUVmax). TBR for SUVmean and SUVmax were higher for 68Ga-Pentixafor than for 18F-FET (SUVmean 154.0±90.7 vs. 4.1±1.3; SUVmax 70.3±44.0 and 3.8±1.2, p<0.01), respectively. Histological analysis confirmed CXCR4 expression in tumor areas with high 68Ga-Pentixafor uptake; regions of the same tumor without apparent 68Ga-Pentixafor uptake showed no or low receptor expression. In this pilot study, 68Ga-Pentixafor retention has been observed in the vast majority of glioblastoma lesions and served as readout for non-invasive determination of CXCR4 expression. Given the paramount importance of the CXCR4/SDF-1 axis in tumor biology, 68Ga-Pentixafor-PET/CT might prove a useful tool for sensitive, non-invasive in-vivo quantification of CXCR4 as well as selection of patients who might benefit from CXCR4-directed therapy. PMID:26909116

  11. Single-cell real-time imaging of transgene expression upon lipofection.

    PubMed

    Fiume, Giuseppe; Di Rienzo, Carmine; Marchetti, Laura; Pozzi, Daniela; Caracciolo, Giulio; Cardarelli, Francesco

    2016-05-20

    Here we address the process of lipofection by quantifying the expression of a genetically-encoded fluorescent reporter at the single-cell level, and in real-time, by confocal imaging in live cells. The Lipofectamine gold-standard formulation is compared to the alternative promising DC-Chol/DOPE formulation. In both cases, we report that only dividing cells are able to produce a detectable amount of the fluorescent reporter protein. Notably, by measuring fluorescence over time in each pair of daughter cells, we find that Lipofectamine-based transfection statistically yields a remarkably higher degree of "symmetry" in protein expression between daughter cells as compared to DC-Chol/DOPE. A model is envisioned in which the degree of symmetry of protein expression is linked to the number of bioavailable DNA copies within the cell before nuclear breakdown. Reported results open new perspectives for the understanding of the lipofection mechanism and define a new experimental platform for the quantitative comparison of transfection reagents. PMID:27012199

  12. PATZ1 is a target of miR-29b that is induced by Ha-Ras oncogene in rat thyroid cells

    PubMed Central

    Vitiello, Michela; Valentino, Teresa; De Menna, Marta; Crescenzi, Elvira; Francesca, Paola; Rea, Domenica; Arra, Claudio; Fusco, Alfredo; De Vita, Gabriella; Cerchia, Laura; Fedele, Monica

    2016-01-01

    The regulatory transcriptional factor PATZ1 is constantly downregulated in human thyroid cancer where it acts as a tumour suppressor by targeting p53-dependent genes involved in Epithelial-Mesenchymal Transition and cell migration. The aim of the present work was to elucidate the upstream signalling mechanisms regulating PATZ1 expression in thyroid cancer cells. The bioinformatics search for microRNAs able to potentially target PATZ1 led to the identification of several miRNAs. Among them we focused on the miR-29b since it was found upregulated in rat thyroid differentiated cells transformed by the Ha-Ras oncogene towards a high proliferating and high migratory phenotype resembling that of anaplastic carcinomas. Functional assays confirmed PATZ1 as a target of miR-29b, and, consistently, an inverse correlation between miR-29b and PATZ1 protein levels was found upon induction of Ha-Ras oncogene expression in these cells. Interestingly, restoration of PATZ1 expression in rat thyroid cells stably expressing the Ha-Ras oncogene decreased cell proliferation and migration, indicating a key role of PATZ1 in Ras-driven thyroid transformation. Together, these results suggest a novel mechanism regulating PATZ1 expression based on the upregulation of miR-29b expression induced by Ras oncogene. PMID:27125250

  13. PATZ1 is a target of miR-29b that is induced by Ha-Ras oncogene in rat thyroid cells.

    PubMed

    Vitiello, Michela; Valentino, Teresa; De Menna, Marta; Crescenzi, Elvira; Francesca, Paola; Rea, Domenica; Arra, Claudio; Fusco, Alfredo; De Vita, Gabriella; Cerchia, Laura; Fedele, Monica

    2016-01-01

    The regulatory transcriptional factor PATZ1 is constantly downregulated in human thyroid cancer where it acts as a tumour suppressor by targeting p53-dependent genes involved in Epithelial-Mesenchymal Transition and cell migration. The aim of the present work was to elucidate the upstream signalling mechanisms regulating PATZ1 expression in thyroid cancer cells. The bioinformatics search for microRNAs able to potentially target PATZ1 led to the identification of several miRNAs. Among them we focused on the miR-29b since it was found upregulated in rat thyroid differentiated cells transformed by the Ha-Ras oncogene towards a high proliferating and high migratory phenotype resembling that of anaplastic carcinomas. Functional assays confirmed PATZ1 as a target of miR-29b, and, consistently, an inverse correlation between miR-29b and PATZ1 protein levels was found upon induction of Ha-Ras oncogene expression in these cells. Interestingly, restoration of PATZ1 expression in rat thyroid cells stably expressing the Ha-Ras oncogene decreased cell proliferation and migration, indicating a key role of PATZ1 in Ras-driven thyroid transformation. Together, these results suggest a novel mechanism regulating PATZ1 expression based on the upregulation of miR-29b expression induced by Ras oncogene. PMID:27125250

  14. Temporal Heterogeneity of Estrogen Receptor Expression in Bone-Dominant Breast Cancer: 18F-Fluoroestradiol PET Imaging Shows Return of ER Expression

    PubMed Central

    Currin, Erin; Peterson, Lanell M.; Schubert, Erin K.; Link, Jeanne M.; Krohn, Kenneth A.; Livingston, Robert B.; Mankoff, David A.; Linden, Hannah M.

    2016-01-01

    Changes in estrogen receptor (ER) expression over the course of therapy may affect response to endocrine therapy. However, measuring temporal changes in ER expression requires serial biopsies, which are impractical and poorly tolerated by most patients. Functional ER imaging using 18F-fluoroestradiol (FES)-PET provides a noninvasive measure of regional ER expression and is ideally suited to serial studies. Additionally, lack of measurable FES uptake in metastatic sites of disease predict tumor progression in patients with ER-positive primary tumors treated with endocrine therapy. This report presents a case of restored sensitivity to endocrine therapy in a patient with bone-dominant breast cancer who underwent serial observational FES-PET imaging over the course of several treatments at our center, demonstrating the temporal heterogeneity of regional ER expression. Although loss and restoration of endocrine sensitivity in patients who have undergone prior hormonal and cytotoxic treatments has been reported, this is, to our knowledge, the first time the accompanying changes in ER expression have been documented by molecular imaging. PMID:26850484

  15. Identification of Novel Small Molecule Inhibitors of Oncogenic RET Kinase

    PubMed Central

    Moccia, Marialuisa; Liu, Qingsong; Guida, Teresa; Federico, Giorgia; Brescia, Annalisa; Zhao, Zheng; Choi, Hwan Geun; Deng, Xianming; Tan, Li; Wang, Jinhua; Billaud, Marc; Gray, Nathanael S.

    2015-01-01

    Oncogenic mutation of the RET receptor tyrosine kinase is observed in several human malignancies. Here, we describe three novel type II RET tyrosine kinase inhibitors (TKI), ALW-II-41-27, XMD15-44 and HG-6-63-01, that inhibit the cellular activity of oncogenic RET mutants at two digit nanomolar concentration. These three compounds shared a 3-trifluoromethyl-4-methylpiperazinephenyl pharmacophore that stabilizes the ‘DFG-out’ inactive conformation of RET activation loop. They blocked RET-mediated signaling and proliferation with an IC50 in the nM range in fibroblasts transformed by the RET/C634R and RET/M918T oncogenes. They also inhibited autophosphorylation of several additional oncogenic RET-derived point mutants and chimeric oncogenes. At a concentration of 10 nM, ALW-II-41-27, XMD15-44 and HG-6-63-01 inhibited RET kinase and signaling in human thyroid cancer cell lines carrying oncogenic RET alleles; they also inhibited proliferation of cancer, but not non-tumoral Nthy-ori-3-1, thyroid cells, with an IC50 in the nM range. The three compounds were capable of inhibiting the ‘gatekeeper’ V804M mutant which confers substantial resistance to established RET inhibitors. In conclusion, we have identified a type II TKI scaffold, shared by ALW-II-41-27, XMD15-44 and HG-6-63-01, that may be used as novel lead for the development of novel agents for the treatment of cancers harboring oncogenic activation of RET. PMID:26046350

  16. SPECT imaging of neuropilin receptor type-1 expression with 131I-labeled monoclonal antibody.

    PubMed

    Dou, Xiaofeng; Yan, Jianghua; Zhang, Yafei; Liu, Peng; Jiang, Yizhen; Lv, Sha; Zeng, Fanwei; Chen, Xiaoli; Wang, Shengyu; Zhang, Haipeng; Wu, Hua; Zhang, Hong; Ouyang, Lin; Su, Xinhui

    2016-09-01

    As a novel co-receptor for vascular endothelial growth factor (VEGF), neuropilin receptor type-1 (NRP-1) is overexpressed in several cancers and metastases, and serves as an attractive target for cancer molecular imaging and therapy. Previous single photon emission computerized tomography (SPECT) studies demonstrated that the small NRP-1-targeting peptides 99mTc-MA-ATWLPPR and 99mTc-CK3 showed poor tumor imaging quality, because of their rapid blood clearance and very low tumor uptake. Compared with small peptides, monoclonal antibodies (mAbs) can improve imaging of NRP-1-expression, due to their high affinity, specificity and slow extraction. A6-11-26 is a novel monoclonal antibody against NRP-1 b1b2 domain that exhibits inhibition of tumor growth in NPR-1-expressing preclinical models. The aim of the present study was to develop the 131I-labeled anti-NRP-1 monoclonal antibody A6-11-26 as a SPECT probe for imaging of NRP-1-positive tumor. An anti-NRP-1 monoclonal antibody (A6-11-26) was produced by hybridomas and was labeled with iodine-131 by the iodogen method. In vitro, the radiolabeling efficiency, radiochemical purity, immunoreactive fraction and stability were assessed. Binding affinity and specificity of 131I‑A6-11-26 to NRP-1 were evaluated using human glioblastoma U87MG cells. In vivo, biodistribution and SPECT/CT studies were conducted on mice bearing U87MG xenografts after the injection of 131I-A6-11-26 with or without co-injection of unlabeled A6-11-26 antibody. A6-11-26 was generated successfully by hybridoma with high purity (>95%) and was labeled with iodine-131 within 60 min with high labelling efficiency (95.46±3.34%), radiochemical purity (98.23±1.41%). 131I-A6-11-26 retained its immunoreactivity and also displayed excellent stability in mouse serum and PBS solution during 1 to 96 h. Cell uptake assays showed high NRP-1-specific uptake (15.80±1.30% applied activity at 6 h) in U87MG cells. 131I-A6-11-26 bound to NRP-1 with low nanomolar

  17. SU-E-I-81: Targeting of HER2-Expressing Tumors with Dual PET-MR Imaging Probes

    SciTech Connect

    Xu, P; Peng, Y; Sun, M; Yang, X

    2015-06-15

    Purpose: The detection of human epidermal growth factor receptor type 2 (HER2) expression in malignant tumors provides important information influencing patient management. Radionuclide in vivo imaging of HER2 may permit the detection of HER2 in both primary tumors and metastases by a single noninvasive procedure. Trastuzumab, effective in about 15 % of women with breast cancer, downregulates signalling through the Akt/PI3K and MAPK pathways.These pathways modulate metabolism which can be monitored by positron emission tomography (PET) and magnetic resonance imaging (MRI). Methods: The relationship between response of HER2 overexpressing tumours and changes in imaging PET or SPECT and MRI will be examined by a integrated bimodal imaging probe.Small (7 kDa) high-affinity anti-HER2 Affibody molecules and KCCYSL targeting peptide may be suitable tracers for visualization of HER2-expressing tumors. Peptide-conjugated iron oxide nanoparticles (Fe3O4 NPs) as MRI imaging and CB-TE2A as PET imaging are integrated into a single synthetic molecule in the HER2 positive cancer. Results: One of targeted contrast bimodal imaging probe agents was synthesized and evaluated to target HER2-expressing tumors in a HER2 positive rat model. We will report the newest results regarding the development of bimodal imaging probes. Conclusion: The preliminary results of the bimodal imaging probe presents high correlation of MRI signal and PET imaging intensity in vivo. This unique feature can hardly be obtained by single model contrast agents. It is envisioned that this bimodal agents can hold great potential for accurate detection of HER2-expressing tumors which are critical for clinical management of the disease.

  18. The HRSC Experiment on Mars Express: First Imaging Results from the Commissioning Phase

    NASA Astrophysics Data System (ADS)

    Oberst, J.; Neukum, G.; Hoffmann, H.; Jaumann, R.; Hauber, E.; Albertz, J.; McCord, T. B.; Markiewicz, W. J.

    2004-12-01

    The ESA Mars Express spacecraft was launched from Baikonur on June 2, 2003, entered Mars orbit on December 25, 2003, and reached the nominal mapping orbit on January 28, 2004. Observing conditions were favorable early on for the HRSC (High Resolution Stereo Camera), designed for the mapping of the Martian surface in 3-D. The HRSC is a pushbroom scanner with 9 CCD line detectors mounted in parallel and perpendicular to the direction of flight on the focal plane. The camera can obtain images at high resolution (10 m/pix), in triple stereo (20 m/pix), in four colors, and at five different phase angles near-simultaneously. An additional Super-Resolution Channel (SRC) yields nested-in images at 2.3 m/pix for detailed photogeologic studies. Even for nominal spacecraft trajectory and camera pointing data from the commissioning phase, solid stereo image reconstructions are feasible. More yet, the three-line stereo data allow us to identify and correct errors in navigation data. We find that > 99% of the stereo rays intersect within a sphere of radius < 20m after orbit and pointing data correction. From the HRSC images we have produced Digital Terrain Models (DTMs) with pixel sizes of 200 m, some of them better. HRSC stereo models and data obtained by the MOLA (Mars Orbiting Laser Altimeter) show good qualitative agreement. Differences in absolute elevations are within 50 m, but may reach several 100 m in lateral positioning (mostly in the spacecraft along-track direction). After correction of these offsets, the HRSC topographic data conveniently fill the gaps between the MOLA tracks and reveal hitherto unrecognized morphologic detail. At the time of writing, the HRSC has covered approx. 22.5 million square kilometers of the Martian surface. In addition, data from 5 Phobos flybys from May through August 2004 were obtained. The HRSC is beginning to make major contributions to geoscience, atmospheric science, photogrammetry, and cartography of Mars (papers submitted to Nature).

  19. Intravital Microscopy for Imaging Subcellular Structures in Live Mice Expressing Fluorescent Proteins

    PubMed Central

    Masedunskas, Andrius; Porat-Shliom, Natalie; Tora, Muhibullah; Milberg, Oleg; Weigert, Roberto

    2013-01-01

    Here we describe a procedure to image subcellular structures in live rodents that is based on the use of confocal intravital microscopy. As a model organ, we use the salivary glands of live mice since they provide several advantages. First, they can be easily exposed to enable access to the optics, and stabilized to facilitate the reduction of the motion artifacts due to heartbeat and respiration. This significantly facilitates imaging and tracking small subcellular structures. Second, most of the cell populations of the salivary glands are accessible from the surface of the organ. This permits the use of confocal microscopy that has a higher spatial resolution than other techniques that have been used for in vivo imaging, such as two-photon microscopy. Finally, salivary glands can be easily manipulated pharmacologically and genetically, thus providing a robust system to investigate biological processes at a molecular level. In this study we focus on a protocol designed to follow the kinetics of the exocytosis of secretory granules in acinar cells and the dynamics of the apical plasma membrane where the secretory granules fuse upon stimulation of the beta-adrenergic receptors. Specifically, we used a transgenic mouse that co-expresses cytosolic GFP and a membrane-targeted peptide fused with the fluorescent protein tandem-Tomato. However, the procedures that we used to stabilize and image the salivary glands can be extended to other mouse models and coupled to other approaches to label in vivo cellular components, enabling the visualization of various subcellular structures, such as endosomes, lysosomes, mitochondria, and the actin cytoskeleton. PMID:24022089

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

    SciTech Connect

    Coppé, 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.

  1. Intrinsically active variants of Erk oncogenically transform cells and disclose unexpected autophosphorylation capability that is independent of TEY phosphorylation

    PubMed Central

    Smorodinsky-Atias, Karina; Goshen-Lago, Tal; Goldberg-Carp, Anat; Melamed, Dganit; Shir, Alexei; Mooshayef, Navit; Beenstock, Jonah; Karamansha, Yael; Darlyuk-Saadon, Ilona; Livnah, Oded; Ahn, Natalie G.; Admon, Arie; Engelberg, David

    2016-01-01

    The receptor-tyrosine kinase (RTK)/Ras/Raf pathway is an essential cascade for mediating growth factor signaling. It is abnormally overactive in almost all human cancers. The downstream targets of the pathway are members of the extracellular regulated kinases (Erk1/2) family, suggesting that this family is a mediator of the oncogenic capability of the cascade. Although all oncogenic mutations in the pathway result in strong activation of Erks, activating mutations in Erks themselves were not reported in cancers. Here we used spontaneously active Erk variants to check whether Erk’s activity per se is sufficient for oncogenic transformation. We show that Erk1(R84S) is an oncoprotein, as NIH3T3 cells that express it form foci in tissue culture plates, colonies in soft agar, and tumors in nude mice. We further show that Erk1(R84S) and Erk2(R65S) are intrinsically active due to an unusual autophosphorylation activity they acquire. They autophosphorylate the activatory TEY motif and also other residues, including the critical residue Thr-207 (in Erk1)/Thr-188 (in Erk2). Strikingly, Erk2(R65S) efficiently autophosphorylates its Thr-188 even when dually mutated in the TEY motif. Thus this study shows that Erk1 can be considered a proto-oncogene and that Erk molecules possess unusual autoregulatory properties, some of them independent of TEY phosphorylation. PMID:26658610

  2. Ultrasound Molecular Imaging of Vascular Endothelial Growth Factor Receptor 2 Expression for Endometrial Receptivity Evaluation

    PubMed Central

    Liu, Hongmei; Chen, Yihan; Yan, Fei; Han, Xiaohua; Wu, Junru; Liu, Xin; Zheng, Hairong

    2015-01-01

    Purpose: Ultrasound (US) molecular imaging by examining the expression of vascular endothelial growth factor receptor 2 (VEGFR2) on uterus vascular endothelium was applied to evaluate the endometrial receptivity. Methods: VEGFR2-targeted ultrasound contrast agents (UCA) and the control UCA (without VEGFR2) were prepared and characterized. Adhesion experiment in vitro was performed with mouse microvascular endothelial cells (bEnd.3) and the ratio of the number of UCA to that of cells at the same field was compared. In vivo study, randomized boluses of targeted or control UCA were injected into the animals of non-pregnancy (D0), pregnancy on day 2 (D2) and day 4 (D4), respectively. Sonograms were acquired by an ultrasound equipment with a 40-MHz high-frequency transducer (Vevo 2100; VisualSonics, Toronto, Canada). The ultrasonic imaging signals were quantified as the video intensity amplitudes generated by the attachment of VEGFR2-targeted UCA. Immunoblotting and immunofluorescence assays were used for confirmation of VEGFR2 expression. Results: Our results showed that VEGFR2-targeted UCA could bind to bEnd.3 cells with significantly higher affinity than the control UCA (9.8 ± 1.0 bubbles/cell versus 0.7 ± 0.3 bubbles/cell, P < 0.01) in vitro. The mean video intensity from the US backscattering of the retained VEGFR2-targeted UCA was significantly higher than that of the control UCA in D2 and D4 mice (D2, 10.5 ± 2.5 dB versus 1.5 ± 1.1 dB, P < 0.01; D4, 15.7 ± 4.0 dB versus 1.5 ± 1.2 dB, P < 0.01), but not significantly different in D0 mice (1.0 ± 0.8 dB versus 0.9 ± 0.6 dB, P > 0.05). Moreover, D4 mice showed the highest video intensity amplitude, indicating the highest VEGFR2 expression when compared with D2 and D0 mice (P < 0.01). This was further confirmed by our immunoblotting and immunofluorescence experiments. Conclusion: Ultrasound molecular imaging with VEGFR2-targeted UCA may be used for noninvasive evaluation of endometrial receptivity in murine

  3. The dark and the bright side of Stat3: proto-oncogene and tumor-suppressor.

    PubMed

    Ecker, Andrea; Simma, Olivia; Hoelbl, Andrea; Kenner, Lukas; Beug, Hartmut; Moriggl, Richard; Sexl, Veronika

    2009-01-01

    Stat transcription factors have been implicated in tumorigenesis in mice and men. Stat3 and Stat5 are considered powerful proto-oncogenes, whereas Stat1 has been demonstrated to suppress tumor formation. We demonstrate here for the first time that a constitutive active version of Stat3alpha (Stat3alphaC) may also suppress transformation. Mouse embryonic fibroblasts (MEFs) deficient for p53 can be transformed with either c-myc or with rasV12 alone. Interestingly, transformation by c-myc is efficiently suppressed by co-expression of Stat3alphaC, but Stat3alphaC does not interfere with transformation by the rasV12-oncogene. In contrast, transplantation of bone marrow cells expressing Stat3alphaC induces the formation of a highly aggressive T cell leukemia in mice. The leukemic cells invaded multiple organs including lung, heart, salivary glands, liver and kidney. Interestingly, transplanted mice developed a similar leukemia when the bone marrow cells were transduced with Stat3beta, which is also constitutively active when expressed at significant levels. Our experiments demonstrate that Stat3 has both - tumor suppressing and tumor promoting properties. PMID:19273247

  4. The FBI1/Akirin2 Target Gene, BCAM, Acts as a Suppressive Oncogene

    PubMed Central

    Akiyama, Hirotada; Iwahana, Yoshimasa; Suda, Mikiya; Yoshimura, Atsunori; Kogai, Hiroyuki; Nagashima, Ai; Ohtsuka, Hiroko; Komiya, Yuko; Tashiro, Fumio

    2013-01-01

    Basal cell adhesion molecule (BCAM), known to be a splicing variant of Lutheran glycoprotein (LU), is an immunoglobulin superfamily membrane protein that acts as a laminin α5 receptor. The high affinity of BCAM/LU for laminin α5 is thought to contribute to the pathogenesis of sickle red blood cells and to various developmental processes. However, the function of BCAM in carcinogenesis is poorly understood. Based on microarray expression analysis, we found that BCAM was one of the target genes of the oncogenic 14-3-3β-FBI1/Akirin2 complex, which acts as a transcriptional repressor and suppresses MAPK phosphatase-1 gene expression. To elucidate the detailed function of BCAM in malignant tumors, we established BCAM-expressing hepatoma K2 cells. These cells lost the malignant characteristics of parental cells, such as anchorage-independent growth, migration, invasion, and tumorigenicity. Moreover, luciferase reporter assays and chromatin immunoprecipitation analysis revealed that the 14-3-3β-FBI1/Akirin2 complex bound to the BCAM promoter and repressed transcription. Thus, these data indicate that BCAM is a suppressive oncoprotein, and that FBI1/Akirin2 is involved in tumorigenicity and metastasis of hepatoma through the downregulation of suppressive oncogenes. PMID:24223164

  5. The FBI1/Akirin2 target gene, BCAM, acts as a suppressive oncogene.

    PubMed

    Akiyama, Hirotada; Iwahana, Yoshimasa; Suda, Mikiya; Yoshimura, Atsunori; Kogai, Hiroyuki; Nagashima, Ai; Ohtsuka, Hiroko; Komiya, Yuko; Tashiro, Fumio

    2013-01-01

    Basal cell adhesion molecule (BCAM), known to be a splicing variant of Lutheran glycoprotein (LU), is an immunoglobulin superfamily membrane protein that acts as a laminin α5 receptor. The high affinity of BCAM/LU for laminin α5 is thought to contribute to the pathogenesis of sickle red blood cells and to various developmental processes. However, the function of BCAM in carcinogenesis is poorly understood. Based on microarray expression analysis, we found that BCAM was one of the target genes of the oncogenic 14-3-3β-FBI1/Akirin2 complex, which acts as a transcriptional repressor and suppresses MAPK phosphatase-1 gene expression. To elucidate the detailed function of BCAM in malignant tumors, we established BCAM-expressing hepatoma K2 cells. These cells lost the malignant characteristics of parental cells, such as anchorage-independent growth, migration, invasion, and tumorigenicity. Moreover, luciferase reporter assays and chromatin immunoprecipitation analysis revealed that the 14-3-3β-FBI1/Akirin2 complex bound to the BCAM promoter and repressed transcription. Thus, these data indicate that BCAM is a suppressive oncoprotein, and that FBI1/Akirin2 is involved in tumorigenicity and metastasis of hepatoma through the downregulation of suppressive oncogenes. PMID:24223164

  6. Oncogenic transformation of Drosophila somatic cells induces a functional piRNA pathway.

    PubMed

    Fagegaltier, Delphine; Falciatori, Ilaria; Czech, Benjamin; Castel, Stephane; Perrimon, Norbert; Simcox, Amanda; Hannon, Gregory J

    2016-07-15

    Germline genes often become re-expressed in soma-derived human cancers as "cancer/testis antigens" (CTAs), and piRNA (PIWI-interacting RNA) pathway proteins are found among CTAs. However, whether and how the piRNA pathway contributes to oncogenesis in human neoplasms remain poorly understood. We found that oncogenic Ras combined with loss of the Hippo tumor suppressor pathway reactivates a primary piRNA pathway in Drosophila somatic cells coincident with oncogenic transformation. In these cells, Piwi becomes loaded with piRNAs derived from annotated generative loci, which are normally restricted to either the germline or the somatic follicle cells. Negating the pathway leads to increases in the expression of a wide variety of transposons and also altered expression of some protein-coding genes. This correlates with a reduction in the proliferation of the transformed cells in culture, suggesting that, at least in this context, the piRNA pathway may play a functional role in cancer. PMID:27474441

  7. Scribble acts as an oncogene in Eμ-myc-driven lymphoma.

    PubMed

    Hawkins, E D; Oliaro, J; Ramsbottom, K M; Newbold, A; Humbert, P O; Johnstone, R W; Russell, S M

    2016-03-01

    Scribble complex proteins maintain apicobasal polarity, regulate cell fate determination and function as tumour suppressors in epithelial tissue. Despite evidence that the function of Scribble is maintained in the lymphocyte lineage, we still understand little about its role as a tumour suppressor in haematological malignancies. Using the Eμ-myc model of Burkitt's lymphoma we investigated the role of Scribble in lymphomagenesis. We found that contrary to its well-documented tumour suppressor role in epithelial tissue, loss of Scribble expression delayed the expansion of peripheral B cells and delayed the onset of Eμ-myc-driven lymphoma. This was despite upregulated ERK phosphorylation levels in Scribble-deficient tumours, which are associated with loss of Scribble expression and the development of more aggressive Burkitt's lymphoma. Interestingly, the developmental stage of lymphoma was unaffected by Scribble expression challenging any role for Scribble in fate determination in the haematopoetic lineage. These data provide evidence for oncogenic properties of Scribble in Myc-driven B-cell lymphomagenesis, reinforcing recent findings that overexpression of a mutant form of Scribble can act as an oncogene in epithelial cells. Our results support the growing appreciation that the tumour regulatory functions of Scribble, and other polarity protein family members, are context dependent. PMID:25982280

  8. Antineoplastic Effects of siRNA against TMPRSS2-ERG Junction Oncogene in Prostate Cancer.

    PubMed

    Urbinati, Giorgia; Ali, Hafiz Muhammad; Rousseau, Quentin; Chapuis, Hubert; Desmaële, Didier; Couvreur, Patrick; Massaad-Massade, Liliane

    2015-01-01

    TMPRSS2-ERG junction oncogene is present in more than 50% of patients with prostate cancer and its expression is frequently associated with poor prognosis. Our aim is to achieve gene knockdown by siRNA TMPRSS2-ERG and then to assess the biological consequences of this inhibition. First, we designed siRNAs against the two TMPRSS2-ERG fusion variants (III and IV), most frequently identified in patients' biopsies. Two of the five siRNAs tested were found to efficiently inhibit mRNA of both TMPRSS2-ERG variants and to decrease ERG protein expression. Microarray analysis further confirmed ERG inhibition by both siRNAs TMPRSS2-ERG and revealed one common down-regulated gene, ADRA2A, involved in cell proliferation and migration. The siRNA against TMPRSS2-ERG fusion variant IV showed the highest anti-proliferative effects: Significantly decreased cell viability, increased cleaved caspase-3 and inhibited a cluster of anti-apoptotic proteins. To propose a concrete therapeutic approach, siRNA TMPRSS2-ERG IV was conjugated to squalene, which can self-organize as nanoparticles in water. The nanoparticles of siRNA TMPRSS2-ERG-squalene injected intravenously in SCID mice reduced growth of VCaP xenografted tumours, inhibited oncoprotein expression and partially restored differentiation (decrease in Ki67). In conclusion, this study offers a new prospect of treatment for prostate cancer based on siRNA-squalene nanoparticles targeting TMPRSS2-ERG junction oncogene. PMID:25933120

  9. Expression of metalloproteinases endometrial stromal sarcoma: immunohistochemical study using image analysis.

    PubMed Central

    Liokumovich, P; Goldberg, I; Davidson, B; Gotlieb, W H; Zahavi, T; Ben-Baruch, G; Reder, I; Kopolovic, J

    1999-01-01

    AIM: To investigate the expression of matrix metalloproteinases (MMP), a group of proteolytic enzymes with a central role in extracellular matrix invasion and degradation, in stromal sarcomas. METHODS: 11 endometrial stromal sarcomas (four low grade tumours, seven high grade) were stained for MMP-2, MMP-3, and MMP-9 using immunohistochemical stains. The surgical material consisted of nine hysterectomy specimens and two pelvic recurrences. Three hysterectomy specimens, removed for leiomyomas, were studied as controls. Staining area was evaluated using image analysis. RESULTS: Age at the time of diagnosis ranged from 21 to 67 years. Four of the 11 patients (three with high grade tumours and one with a low grade tumour) died of the disease, six remained free of disease, and one was lost to follow up. Staining for MMP-2, MMP-3, and MMP-9 was more diffuse in high grade tumours than in low grade tumours and controls. Staining for MMP-3 and MMP-9 was more pronounced in high grade than in low grade tumours (p = 0.04; p = 0.05). Staining for MMP-9 was significantly greater in all stromal sarcomas than in controls (p < 0.001 for high grade tumours v controls; p < 0.01 for low grade tumours v controls). Diffuse staining for MMP-2, exceeding 90% of the tumour area, was observed in three of seven high grade tumours but in no low grade tumours. There was no apparent correlation between staining for any of the three enzymes and survival. CONCLUSIONS: Both low and high grade endometrial stromal tumours express matrix metalloproteinases. MMP-3 and MMP-9 are expressed more diffusely in high grade than in low grade tumours. In the individual case, diffuse staining for MMP-2 appears to best characterise the high grade tumours. Thus staining for MMP-2 may aid in differentiating high grade from low grade tumours, and MMP-9 in differentiating normal endometrial stroma from low and high grade endometrial stromal sarcomas. MMP expression does not appear to predict disease outcome in

  10. Analyzing in situ gene expression in the mouse brain with image registration, feature extraction and block clustering

    PubMed Central

    Jagalur, Manjunatha; Pal, Chris; Learned-Miller, Erik; Zoeller, R Thomas; Kulp, David

    2007-01-01

    Background Many important high throughput projects use in situ hybridization and may require the analysis of images of spatial cross sections of organisms taken with cellular level resolution. Projects creating gene expression atlases at unprecedented scales for the embryonic fruit fly as well as the embryonic and adult mouse already involve the analysis of hundreds of thousands of high resolution experimental images mapping mRNA expression patterns. Challenges include accurate registration of highly deformed tissues, associating cells with known anatomical regions, and identifying groups of genes whose expression is coordinately regulated with respect to both concentration and spatial location. Solutions to these and other challenges will lead to a richer understanding of the complex system aspects of gene regulation in heterogeneous tissue. Results We present an end-to-end approach for processing raw in situ expression imagery and performing subsequent analysis. We use a non-linear, information theoretic based image registration technique specifically adapted for mapping expression images to anatomical annotations and a method for extracting expression information within an anatomical region. Our method consists of coarse registration, fine registration, and expression feature extraction steps. From this we obtain a matrix for expression characteristics with rows corresponding to genes and columns corresponding to anatomical sub-structures. We perform matrix block cluster analysis using a novel row-column mixture model and we relate clustered patterns to Gene Ontology (GO) annotations. Conclusion Resulting registrations suggest that our method is robust over intensity levels and shape variations in ISH imagery. Functional enrichment studies from both simple analysis and block clustering indicate that gene relationships consistent with biological knowledge of neuronal gene functions can be extracted from large ISH image databases such as the Allen Brain Atlas [1

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

    SciTech Connect

    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.

  12. Inhibition of ras oncogene: a novel approach to antineoplastic therapy.

    PubMed

    Scharovsky, O G; Rozados, V R; Gervasoni, S I; Matar, P

    2000-01-01

    The most frequently detected oncogene alterations, both in animal and human cancers, are the mutations in the ras oncogene family. These oncogenes are mutated or overexpressed in many human tumors, with a high incidence in tumors of the pancreas, thyroid, colon, lung and certain types of leukemia. Ras is a small guanine nucleotide binding protein that transduces biological information from the cell surface to cytoplasmic components within cells. The signal is transduced to the cell nucleus through second messengers, and it ultimately induces cell division. Oncogenic forms of p21(ras) lead to unregulated, sustained signaling through downstream effectors. The ras family of oncogenes is involved in the development of both primary tumors and metastases making it a good therapeutic target. Several therapeutic approaches to cancer have been developed pointing to reducing the altered gene product or to eliminating its biological function: (1) gene therapy with ribozymes, which are able to break down specific RNA sequences, or with antisense oligonucleotides, (2) immunotherapy through passive or active immunization protocols, and (3) inhibition of p21(ras) farnesylation either by inhibition of farnesyl transferase or synthesis inhibition of farnesyl moieties. PMID:10895051

  13. The transcription factor LSF: a novel oncogene for hepatocellular carcinoma

    PubMed Central

    Santhekadur, Prasanna K; Rajasekaran, Devaraja; Siddiq, Ayesha; Gredler, Rachel; Chen, Dong; Schaus, Scott E; Hansen, Ulla; Fisher, Paul B; Sarkar, Devanand

    2012-01-01

    The transcription factor LSF (Late SV40 Factor), also known as TFCP2, belongs to the LSF/CP2 family related to Grainyhead family of proteins and is involved in many biological events, including regulation of cellular and viral promoters, cell cycle, DNA synthesis, cell survival and Alzheimer’s disease. Our recent studies establish an oncogenic role of LSF in Hepatocellular carcinoma (HCC). LSF overexpression is detected in human HCC cell lines and in more than 90% cases of human HCC patients, compared to normal hepatocytes and liver, and its expression level showed significant correlation with the stages and grades of the disease. Forced overexpression of LSF in less aggressive HCC cells resulted in highly aggressive, angiogenic and multi-organ metastatic tumors in nude mice. Conversely, inhibition of LSF significantly abrogated growth and metastasis of highly aggressive HCC cells in nude mice. Microarray studies revealed that as a transcription factor LSF modulated specific genes regulating invasion, angiogenesis, chemoresistance and senescence. LSF transcriptionally regulates thymidylate synthase (TS) gene, thus contributing to cell cycle regulation and chemoresistance. Our studies identify a network of proteins, including osteopontin (OPN), Matrix metalloproteinase-9 (MMP-9), c-Met and complement factor H (CFH), that are directly regulated by LSF and play important role in LSF-induced hepatocarcinogenesis. A high throughput screening identified small molecule inhibitors of LSF DNA binding and the prototype of these molecules, Factor Quinolinone inhibitor 1 (FQI1), profoundly inhibited cell viability and induced apoptosis in human HCC cells without exerting harmful effects to normal immortal human hepatocytes and primary mouse hepatocytes. In nude mice xenograft studies, FQI1 markedly inhibited growth of human HCC xenografts as well as angiogenesis without exerting any toxicity. These studies establish a key role of LSF in hepatocarcinogenesis and usher in a

  14. Immunoprevention of Chemical Carcinogenesis through Early Recognition of Oncogene Mutations

    PubMed Central

    Nasti, Tahseen H.; Rudemiller, Kyle J.; Cochran, J. Barry; Kim, Hee Kyung; Tsuruta, Yuko; Fineberg, Naomi S.; Athar, Mohammad

    2015-01-01

    Prevention of tumors induced by environmental carcinogens has not been achieved. Skin tumors produced by polyaromatic hydrocarbons, such as 7,12-dimethylbenz(a)anthracene (DMBA), often harbor an H-ras point mutation, suggesting that it is a poor target for early immunosurveillance. The application of pyrosequencing and allele-specific PCR techniques established that mutations in the genome and expression of the Mut H-ras gene could be detected as early as 1 d after DMBA application. Further, DMBA sensitization raised Mut H-ras epitope–specific CTLs capable of eliminating Mut H-ras+ preneoplastic skin cells, demonstrating that immunosurveillance is normally induced but may be ineffective owing to insufficient effector pool size and/or immunosuppression. To test whether selective pre-expansion of CD8 T cells with specificity for the single Mut H-ras epitope was sufficient for tumor prevention, MHC class I epitope–focused lentivector-infected dendritic cell– and DNA-based vaccines were designed to bias toward CTL rather than regulatory T cell induction. Mut H-ras, but not wild-type H-ras, epitope-focused vaccination generated specific CTLs and inhibited DMBA-induced tumor initiation, growth, and progression in preventative and therapeutic settings. Transferred Mut H-ras–specific effectors induced rapid tumor regression, overcoming established tumor suppression in tumor-bearing mice. These studies support further evaluation of oncogenic mutations for their potential to act as early tumor-specific, immunogenic epitopes in expanding relevant immunosurveillance effectors to block tumor formation, rather than treating established tumors. PMID:25694611

  15. Modern fluorescent proteins and imaging technologies to study gene expression, nuclear localization, and dynamics

    PubMed Central

    Wu, Bin; Piatkevich, Kiryl D; Lionnet, Timothée; Singer, Robert H; Verkhusha, Vladislav V

    2011-01-01

    Recent developments in reagent design can address problems in single cells that were not previously approachable. We have attempted to foresee what will become possible, and the sorts of biological problems that become tractable with these novel reagents. We have focused on the novel fluorescent proteins that allow convenient multiplexing, and provide for a time-dependent analysis of events in single cells. Methods for fluorescently labeling specific molecules, including endogenously expressed proteins and mRNA have progressed and are now commonly used in a variety of organisms. Finally, sensitive microscopic methods have become more routine practice. This article emphasizes that the time is right to coordinate these approaches for a new initiative on single cell imaging of biological molecules. PMID:21242078

  16. ImmunoPET Imaging of CD146 Expression in Malignant Brain Tumors.

    PubMed

    Hernandez, Reinier; Sun, Haiyan; England, Christopher G; Valdovinos, Hector F; Barnhart, Todd E; Yang, Yunan; Cai, Weibo

    2016-07-01

    Recently, the overexpression of CD146 and its potential as a therapeutic target in high-grade gliomas, the most lethal type of brain cancer, was uncovered. In this study, we describe the generation of (89)Zr-Df-YY146, a novel (89)Zr-labeled monoclonal antibody (mAb) for the targeting and quantification of CD146 expression in a mouse model of glioblastoma, using noninvasive immunoPET imaging. YY146, a high affinity anti-CD146 mAb, was conjugated to deferoxamine (Df) for labeling with the long-lived positron emitter (89)Zr (t1/2: 78.4 h). In vitro assays, including flow cytometry, immunofluorescence microscopy, and Western blot, were performed with two glioblastoma cell lines, U87MG and U251, to determine their CD146 expression levels. Also, YY146 and Df-YY146's CD146-binding affinities were compared using flow cytometry. In vivo CD146-targeting of (89)Zr-Df-YY146 was evaluated by sequential PET imaging, in athymic nude mice bearing subcutaneously implanted U87MG or U251 tumors. CD146 blocking, ex vivo biodistribution, and histological studies were carried out to confirm (89)Zr-Df-YY146 specificity, as well as the accuracy of PET data. In vitro studies exposed elevated CD146 expression levels in U87MG cells, but negligible levels in U251 cells. Flow cytometry revealed no differences in affinity between YY146 and Df-YY146. (89)Zr labeling of Df-YY146 proceeded with excellent yield (∼80%), radiochemical purity (>95%), and specific activity (∼44 GBq/μmol). Longitudinal PET revealed prominent and persistent (89)Zr-Df-YY146 uptake in mice bearing U87MG tumors that peaked at 14.00 ± 3.28%ID/g (n = 4), 48 h post injection of the tracer. Conversely, uptake was significantly lower in CD146-negative U251 tumors (5.15 ± 0.99%ID/g, at 48 h p.i.; n = 4; P < 0.05). Uptake in U87MG tumors was effectively blocked in a competitive inhibition experiment, corroborating the CD146 specificity of (89)Zr-Df-YY146. Finally, ex vivo biodistribution validated the accuracy of PET data

  17. The Plasticity of Oncogene Addiction: Implications for Targeted Therapies Directed to Receptor Tyrosine Kinases12

    PubMed Central

    Pillay, Vinochani; Allaf, Layal; Wilding, Alexander L; Donoghue, Jacqui F; Court, Naomi W; Greenall, Steve A; Scott, Andrew M; Johns, Terrance G

    2009-01-01

    A common mutation of the epidermal growth factor receptor (EGFR) in glioblastoma multiforme (GBM) is an extracellular truncation known as the de2-7 EGFR (or EGFRvIII). Hepatocyte growth factor (HGF) is the ligand for the receptor tyrosine kinase (RTK) c-Met, and this signaling axis is often active in GBM. The expression of the HGF/c-Met axis or de2-7 EGFR independently enhances GBMgrowth and invasiveness, particularly through the phosphatidylinositol-3 kinase/pAkt pathway. Using RTK arrays, we show that expression of de2-7 EGFR in U87MG GBM cells leads to the coactivation of several RTKs, including platelet-derived growth factor receptor β and c-Met. A neutralizing antibody to HGF (AMG102) did not inhibit de2-7 EGFR-mediated activation of c-Met, demonstrating that it is ligand-independent. Therapy for parental U87MG xenografts with AMG 102 resulted in significant inhibition of tumor growth, whereas U87MG.Δ2-7 xenografts were profoundly resistant. Treatment of U87MG.Δ2-7 xenografts with panitumumab, an anti-EGFR antibody, only partially inhibited tumor growth as xenografts rapidly reverted to the HGF/c-Met signaling pathway. Cotreatment with panitumumab and AMG 102 prevented this escape leading to significant tumor inhibition through an apoptotic mechanism, consistent with the induction of oncogenic shock. This observation provides a rationale for using panitumumab and AMG 102 in combination for the treatment of GBM patients. These results illustrate that GBM cells can rapidly change the RTK driving their oncogene addiction if the alternate RTK signals through the same downstream pathway. Consequently, inhibition of a dominant oncogene by targeted therapy can alter the hierarchy of RTKs resulting in rapid therapeutic resistance. PMID:19412429

  18. Oncogenic CARMA1 couples NF-κB and β-catenin signaling in diffuse large B-cell lymphomas

    PubMed Central

    Bognar, M K; Vincendeau, M; Erdmann, T; Seeholzer, T; Grau, M; Linnemann, J R; Ruland, J; Scheel, C H; Lenz, P; Ott, G; Lenz, G; Hauck, S M; Krappmann, D

    2016-01-01

    Constitutive activation of the antiapoptotic nuclear factor-κB (NF-κB) signaling pathway is a hallmark of the activated B-cell-like (ABC) subtype of diffuse large B-cell lymphomas (DLBCL). Recurrent oncogenic mutations are found in the scaffold protein CARMA1 (CARD11) that connects B-cell receptor (BCR) signaling to the canonical NF-κB pathway. We asked how far additional downstream processes are activated and contribute to the oncogenic potential of DLBCL-derived CARMA1 mutants. To this end, we expressed oncogenic CARMA1 in the NF-κB negative DLBCL lymphoma cell line BJAB. By a proteomic approach we identified recruitment of β-catenin and its destruction complex consisting of APC, AXIN1, CK1α and GSK3β to oncogenic CARMA1. Recruitment of the β-catenin destruction complex was independent of CARMA1-BCL10-MALT1 complex formation or constitutive NF-κB activation and promoted the stabilization of β-catenin. The β-catenin destruction complex was also recruited to CARMA1 in ABC DLBCL cell lines, which coincided with elevated β-catenin expression. In line, β-catenin was frequently detected in non-GCB DLBCL biopsies that rely on chronic BCR signaling. Increased β-catenin amounts alone were not sufficient to induce classical WNT target gene signatures, but could augment TCF/LEF-dependent transcriptional activation in response to WNT signaling. In conjunction with NF-κB, β-catenin enhanced expression of immunosuppressive interleukin-10 and suppressed antitumoral CCL3, indicating that β-catenin can induce a favorable tumor microenvironment. Thus, parallel activation of NF-κB and β-catenin signaling by gain-of-function mutations in CARMA1 augments WNT stimulation and is required for regulating the expression of distinct NF-κB target genes to trigger cell-intrinsic and extrinsic processes that promote DLBCL lymphomagenesis. PMID:26776161

  19. Imaging Real-Time Gene Expression in Mammalian Cells with Single-Transcript Resolution

    PubMed Central

    Wells, Amber L.; Condeelis, John S.; Singer, Robert H.; Zenklusen, Daniel

    2016-01-01

    The MS2 system provides optimal sensitivity for single-molecule detection in cells. It requires two genetically encoded moieties: a reporter mRNA that contains MS2 binding site (MBS) stem loops and a fluorescent MS2 coat protein (MCP-xFP) that binds to the stem loops with high affinity, thus tagging the mRNA within the cell. This protocol describes transfection of COS-7 cells with reporter RNA (e.g., pRSV-Z-24 MBS-β-actin) and MCP-xFP (e.g., pPolII-MCP-GFP-NLS) plasmids using calcium phosphate precipitation. The reporter mRNA plasmid must be co-transfected with the MCP-xFP-NLS plasmid for simultaneous expression in a cell. The unbound MCP-xFP-NLS is sequestered in the nucleus, leaving only the MCP-xFP-NLS that is bound to the reporter mRNA in the cytoplasm. This provides a high signal-to-noise ratio (SNR) that permits detection of single mRNA molecules. The Delta T Imaging System is used for image acquisition of fluorescent particles in the cells. PMID:21356977

  20. Non-invasive imaging of transgenic GFP expression in neonatal mouse brain

    NASA Astrophysics Data System (ADS)

    Ho, Gideon; Zhang, Chunyan; Zhuo, Lang

    2007-02-01

    Glial fibrillary acidic protein (GFAP) is a traditional biomarker for astrocytes of the central nervous system. In this study, non-invasive in vivo imaging of GFAP-GFP (green fluorescent protein) expression in the brain of neonatal transgenic mice is used as a novel method to investigate the relationship between the expression of the transgene at 0, 2, 4, 6 and 8 hr post-treatment in mice subjected to a single administration of 12 mg/kg of neurotoxin 1-methyl-4(2'-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH 3-MPTP). The GFP elevation was found to peak at 6 hr and lasted to at least 8 hr after the toxin treatment. Histological examination of fixed brain sections using immunohistochemistry (IHC) shows an increase in GFP and GFAP signal from the substantia nigra pars compacta (SNpc) and the hippocampus. The results have provided quantitative fluorescence and qualitative histological evidence for the activation of the GFAP-GFP transgene in astrocytes following neurotoxin 2'-CH 3-MPTP administration, suggesting that the model described here could be used to study neuronal degeneration such as Parkinson's disease and in general, developmental neurotoxicity in live animals.

  1. Imaging of CAIX-expressing xenografts in vivo using 99mTc-HEHEHE-ZCAIX:1 Affibody molecule

    PubMed Central

    HONARVAR, HADIS; GAROUSI, JAVAD; GUNNERIUSSON, ELIN; HÖIDÉN-GUTHENBERG, INGMARIE; ALTAI, MOHAMED; WIDSTRÖM, CHARLES; TOLMACHEV, VLADIMIR; FREJD, FREDRIK Y.

    2015-01-01

    Carbonic anhydrase IX (CAIX) is a transmembrane enzyme involved in regulation of tissue pH balance. In cancer, CAIX expression is associated with tumor hypoxia. CAIX is also overexpressed in renal cell carcinoma and is a molecular target for the therapeutic antibody cG250 (girentuximab). Radionuclide imaging of CAIX expression might be used for identification of patients who may benefit from cG250 therapy and from treatment strategies for hypoxic tumors. Affibody molecules are small (7 kDa) scaffold proteins having a high potential as probes for radionuclide molecular imaging. The aim of the present study was to evaluate feasibility of in vivo imaging of CAIX-expression using radiolabeled Affibody molecules. A histidine-glutamate-histidine-glutamate-histidine-glutamate (HE)3-tag-containing CAIX-binding Affibody molecule (HE)3-ZCAIX:1 was labeled with [99mTc(CO)3]+. Its binding properties were evaluated in vitro using CAIX-expressing SK-RC-52 renal carcinoma cells. 99mTc-(HE)3-ZCAIX:1 was evaluated in NMRI nu/nu mice bearing SK-RC-52 xenografts. The in vivo specificity test confirmed CAIX-mediated tumor targeting. 99mTc-(HE)3-ZCAIX:1 cleared rapidly from blood and normal tissues except for kidneys. At optimal time-point (4 h p.i.), the tumor uptake was 9.7±0.7% ID/g, and tumor-to-blood ratio was 53±10. Experimental imaging of CAIX-expressing SK-RC-52 xenografts at 4 h p.i. provided high contrast images. The use of radioiodine label for ZCAIX:1 enabled the reduction of renal uptake, but resulted in significantly lower tumor uptake and tumor-to-blood ratio. Results of the present study suggest that radiolabeled Affibody molecules are promising probes for imaging of CAIX-expression in vivo. PMID:25434612

  2. Assessment of α-Fetoprotein Targeted HSV1-tk Expression in Hepatocellular Carcinoma with In Vivo Imaging

    PubMed Central

    Park, Ju Hui; Kim, Kwang Il; Lee, Kyo Chul; Lee, Yong Jin; Lee, Tae Sup; Chung, Wee Sup; Lim, Sang Moo

    2015-01-01

    Abstract Tumor-specific enhancer/promoter is applicable for targeting gene expression in tumors and helpful for tumor-targeting imaging and therapy. We aimed to acquire α-fetoprotein (AFP)-producing hepatocellular carcinoma (HCC) specific images using adenovirus containing HSV1-tk gene controlled by AFP enhancer/promoter and evaluate in vivo ganciclovir (GCV)-medicated therapeutic effects on AFP-targeted HSV1-tk expression with 18F-FDG positron emission tomography (PET). Recombinant adenovirus expressing HSV1-tk under AFP enhancer/promoter was produced (AdAFP-TK) and the expression levels were evaluated by RT-PCR and 125I-IVDU uptake. GCV-mediated HSV1-tk cytotoxicity was determined by MTT assay. After the mixture of AdAFP-fLuc and AdAFP-TK was administrated, bioluminescent images (BLIs) and 18F-FHBG PET images were obtained in tumor-bearing mice. In vivo therapeutic effects of AdAFP-TK and GCV in the HuH-7 xenograft model were monitored by 18F-FDG PET. When infected with AdAFP-TK, cell viability in HuH-7 was reduced, but those in HT-29 and SK-Hep-1 were not significantly decreased at any GCV concentration less than 100 μM. AFP-targeted fLuc and HSV1-tk expression were clearly visualized by BLI and 18F-FHBG PET images in AFP-producing HCC, respectively. In vivo GCV-mediated tumor growth inhibition by AFP-targeted HSV1-tk expression was monitored by 18F-FDG PET. Recombinant AdAFP-TK could be applied for AFP-targeted HCC gene therapy and imaging in AFP-producing HCC. PMID:25545853

  3. Analysis of OMEGA/Mars Express hyperspectral images with a linear unmixing model

    NASA Astrophysics Data System (ADS)

    Le Mouelic, S.; Combe, J.-Ph.; Sotin, C.; Le Deit, L.; Gendrin, A.; Mustard, J.; Bibring, J.-P.; Langevin, Y.; Gondet, B.; Pinet, P.

    The OMEGA imaging spectrometer onboard Mars Express has completed a near global coverage of Mars in 352 spectral channels from 0.3 to 5.1 µm at a spatial resolution ranging from 300 m to 4 km. This unprecedented data set provides the opportunity to investigate the mineralogy of the surface of Mars by looking at diagnostic spectral features in the visible and near infrared domains [1]. We have focused our data reduction approach on the linear unmixing strategies. Working on a pixel by pixel basis, we find the best linear combination of a suite of laboratory spectra of pure minerals which match the OMEGA data. A spectrally flat and dark artificial component is introduced to account for shading effects. Similarly, we use two pure positive and negative slopes to account at first order for continuum slope variations linked to scattering, grain size and photometric effects. This approach allows us to draw several conclusions on the overall mineralogy of the observed regions. In particular, the Syrtis Major area appears dominated by a mixing between low and high Calcium pyroxenes in various amounts, with localized exposures of iron-rich olivines. At a global scale, the southern hemisphere appears enriched in both low-Ca and high-Ca pyroxenes. Signatures of iron oxides are detected in the bright regions of the northern hemisphere. These results agree with those obtained with different approaches such as MGM or ratio images [1,2]. The advantages and limits of the unmixing approach applied to OMEGA hyperspectral images will be discussed. References: [1] Bibring et al. (2005), Science, vol. 307, 5715, 1576-1581. [2] Mustard et al., Science (2005), vol. 307, 5715, 1594-1597.

  4. Opposing oncogenic activities of small DNA tumor virus transforming proteins

    PubMed Central

    Chinnadurai, G.

    2011-01-01

    The E1A gene of species C human adenovirus is an intensely investigated model viral oncogene that immortalizes primary cells and mediates oncogenic cell transformation in cooperation with other viral or cellular oncogenes. Investigations using E1A proteins have illuminated important paradigms in cell proliferation and the functions of cellular proteins such as the retinoblastoma protein. Studies with E1A have led to the surprising discovery that E1A also suppresses cell transformation and oncogenesis. Here, I review our current understanding of the transforming and tumor suppressive functions of E1A, and how E1A studies led to the discovery of a related tumor suppressive function in benign human papillomaviruses. The potential role of these opposing functions in viral replication in epithelial cells is also discussed. PMID:21330137

  5. Oncogene addiction: pathways of therapeutic response, resistance, and road maps toward a cure

    PubMed Central

    Pagliarini, Raymond; Shao, Wenlin; Sellers, William R

    2015-01-01

    A key goal of cancer therapeutics is to selectively target the genetic lesions that initiate and maintain cancer cell proliferation and survival. While most cancers harbor multiple oncogenic mutations, a wealth of preclinical and clinical data supports that many cancers are sensitive to inhibition of single oncogenes, a concept referred to as ‘oncogene addiction’. Herein, we describe the clinical evidence supporting oncogene addiction and discuss common mechanistic themes emerging from the response and acquired resistance to oncogene-targeted therapies. Finally, we suggest several opportunities toward exploiting oncogene addiction to achieve curative cancer therapies. PMID:25680965

  6. Metabolic Rewiring by Oncogenic BRAF V600E Links Ketogenesis Pathway to BRAF-MEK1 Signaling.

    PubMed

    Kang, Hee-Bum; Fan, Jun; Lin, Ruiting; Elf, Shannon; Ji, Quanjiang; Zhao, Liang; Jin, Lingtao; Seo, Jae Ho; Shan, Changliang; Arbiser, Jack L; Cohen, Cynthia; Brat, Daniel; Miziorko, Henry M; Kim, Eunhee; Abdel-Wahab, Omar; Merghoub, Taha; Fröhling, Stefan; Scholl, Claudia; Tamayo, Pablo; Barbie, David A; Zhou, Lu; Pollack, Brian P; Fisher, Kevin; Kudchadkar, Ragini R; Lawson, David H; Sica, Gabriel; Rossi, Michael; Lonial, Sagar; Khoury, Hanna J; Khuri, Fadlo R; Lee, Benjamin H; Boggon, Titus J; He, Chuan; Kang, Sumin; Chen, Jing

    2015-08-01

    Many human cancers share similar metabolic alterations, including the Warburg effect. However, it remains unclear whether oncogene-specific metabolic alterations are required for tumor development. Here we demonstrate a "synthetic lethal" interaction between oncogenic BRAF V600E and a ketogenic enzyme 3-hydroxy-3-methylglutaryl-CoA lyase (HMGCL). HMGCL expression is upregulated in BRAF V600E-expressing human primary melanoma and hairy cell leukemia cells. Suppression of HMGCL specifically attenuates proliferation and tumor growth potential of human melanoma cells expressing BRAF V600E. Mechanistically, active BRAF upregulates HMGCL through an octamer transcription factor Oct-1, leading to increased intracellular levels of HMGCL product, acetoacetate, which selectively enhances binding of BRAF V600E but not BRAF wild-type to MEK1 in V600E-positive cancer cells to promote activation of MEK-ERK signaling. These findings reveal a mutation-specific mechanism by which oncogenic BRAF V600E "rewires" metabolic and cell signaling networks and signals through the Oct-1-HMGCL-acetoacetate axis to selectively promote BRAF V600E-dependent tumor development. PMID:26145173

  7. Abnormal structure of the canine oncogene, related to the human c-yes-1 oncogene, in canine mammary tumor tissue.

    PubMed

    Miyoshi, N; Tateyama, S; Ogawa, K; Yamaguchi, R; Kuroda, H; Yasuda, N; Shimizu, T

    1991-12-01

    Cellular oncogenes of genomic DNA in 6 canine primary mammary tumors were screened by Southern blot analysis, using 7 oncogene probes. A canine genomic oncogene related to the human c-yes-1 oncogene was detected as abnormal bands in solid carcinoma genomic DNA digested with EcoRI, HindIII, HindIII-EcoRI, or HindIII-BamHI. Comparison was made between other tumor specimens and control specimens obtained from 4 clinically normal dogs--1 mixed breed and 3 Shiba Inu dogs (the same breed as the dog from which the solid carcinoma was obtained). These abnormal bands were 0.1 to 1 kilobase shorter than the normal gene. However, digestion of genomic DNA obtained from normal WBC of this dog also produced all of the abnormal bands as observed in digested DNA from the solid carcinoma tissue. Therefore, in this dog, the genomic DNA of all somatic cells from the ontogenic stage still had the abnormal sequences related to the human c-yes-1 oncogene, and it is possible that this abnormal structure may have some role (eg, as an initiator) in tumorigenesis or the progression of this tumor. PMID:1789521

  8. Progression of familial adenomatous polyposis (FAP) colonic cells after transfer of the src or polyoma middle T oncogenes: cooperation between src and HGF/Met in invasion.

    PubMed Central

    Empereur, S.; Djelloul, S.; Di Gioia, Y.; Bruyneel, E.; Mareel, M.; Van Hengel, J.; Van Roy, F.; Comoglio, P.; Courtneidge, S.; Paraskeva, C.; Chastre, E.; Gespach, C.

    1997-01-01

    Little is known about the the signalling pathways driving the adenoma-to-carcinoma sequence in human colonic epithelial cells. Accumulation and activation of the src tyrosine kinase in colon cancer suggest a potential role of this oncogene in this early progression. Therefore, we introduced either activated src (m-src), polyoma-MT alone or combined with normal c-src in the adenoma PC/AA/C1 cell line (PC) to define the function and phenotypic transformations induced by these oncogenes in familial adenomatous polyposis (FAP) colonic epithelial cells. Functional expression of these oncoproteins induced the adenoma-to-carcinoma conversion, overexpression of the hepatocyte growth factor (HGF) receptor Met, but failed to confer invasiveness in vivo and in vitro, or to produce alterations in cell proliferation and differentiation. In contrast, PC-msrc cells became susceptible to the HGF-induced invasion of collagen gels and exhibited sustained activation of the pp60src tyrosine kinase and Tyr phosphorylation of the 120-kDa E-cadherin, which was further increased by HGF Transcripts of HGF were clearly identified by reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot in the parental and transformed PC cells, suggesting an autocrine mechanism. Taken together, the data indicate that: (1) experimental activation of src and PyMT pathways directly induces tumorigenicity and Met upregulation in a colon adenoma cell line; (2) HGF-activated Met and src cooperate in inducing invasion; (3) in view of the molecular associations between catenins and cadherin or the tumour-suppressor gene product APC, the cell adhesion molecule E-cadherin may constitute a downstream effector of src and Met. Images Figure 1 Figure 2 Figure 4 Figure 5 Figure 6 PMID:9010033

  9. Bioluminescence Imaging Captures the Expression and Dynamics of Endogenous p21 Promoter Activity in Living Mice and Intact Cells▿

    PubMed Central

    Tinkum, Kelsey L.; Marpegan, Luciano; White, Lynn S.; Sun, Jinwu; Herzog, Erik D.; Piwnica-Worms, David; Piwnica-Worms, Helen

    2011-01-01

    To interrogate endogenous p21WAF1/CIP1 (p21) promoter activity under basal conditions and in response to various forms of stress, knock-in imaging reporter mice in which expression of firefly luciferase (FLuc) was placed under the control of the endogenous p21 promoter within the Cdkn1a gene locus were generated. Bioluminescence imaging (BLI) of p21 promoter activity was performed noninvasively and repetitively in mice and in cells derived from these mice. We demonstrated that expression of FLuc accurately reported endogenous p21 expression at baseline and under conditions of genotoxic stress and that photon flux correlated with mRNA abundance and, therefore, bioluminescence provided a direct readout of p21 promoter activity in vivo. BLI confirmed that p53 was required for activation of the p21 promoter in vivo in response to ionizing radiation. Interestingly, imaging of reporter cells demonstrated that p53 prevents the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway from activating p21 expression when quiescent cells are stimulated with serum to reenter the cell cycle. In addition, low-light BLI identified p21 expression in specific regions of individual organs that had not been observed previously. This inducible p21FLuc knock-in reporter strain will facilitate imaging studies of p53-dependent and -independent stress responses within the physiological context of the whole animal. PMID:21791610

  10. Bioluminescence imaging captures the expression and dynamics of endogenous p21 promoter activity in living mice and intact cells.

    PubMed

    Tinkum, Kelsey L; Marpegan, Luciano; White, Lynn S; Sun, Jinwu; Herzog, Erik D; Piwnica-Worms, David; Piwnica-Worms, Helen

    2011-09-01

    To interrogate endogenous p21(WAF1/CIP1) (p21) promoter activity under basal conditions and in response to various forms of stress, knock-in imaging reporter mice in which expression of firefly luciferase (FLuc) was placed under the control of the endogenous p21 promoter within the Cdkn1a gene locus were generated. Bioluminescence imaging (BLI) of p21 promoter activity was performed noninvasively and repetitively in mice and in cells derived from these mice. We demonstrated that expression of FLuc accurately reported endogenous p21 expression at baseline and under conditions of genotoxic stress and that photon flux correlated with mRNA abundance and, therefore, bioluminescence provided a direct readout of p21 promoter activity in vivo. BLI confirmed that p53 was required for activation of the p21 promoter in vivo in response to ionizing radiation. Interestingly, imaging of reporter cells demonstrated that p53 prevents the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway from activating p21 expression when quiescent cells are stimulated with serum to reenter the cell cycle. In addition, low-light BLI identified p21 expression in specific regions of individual organs that had not been observed previously. This inducible p21(FLuc) knock-in reporter strain will facilitate imaging studies of p53-dependent and -independent stress responses within the physiological context of the whole animal. PMID:21791610

  11. Oncogenic osteomalacia: Problems in diagnosis and long-term management

    PubMed Central

    Dhammi, Ish K; Jain, Anil K; Singh, Ajay Pal; Mishra, Puneet; Jain, Saurabh

    2010-01-01

    Oncogenic osteomalacia is a rare association between mesenchymal tumors and hypophosphatemic rickets. It is more of a biochemical entity than a clinical one. The pathophysiology of the tumor is not clear. However, it has been seen that the clinical and biochemical parameters become normal if the lesion responsible for producing the osteomalacia is excised. For a clinical diagnosis a high index of suspicion is necessary. We present three such cases where in one the oncogenic osteomalacia reversed while in rest it did not. We present this case report to sensitize about the entity. PMID:20924490

  12. Control of c-fos and c-myc proto-oncogene induction in rat thyroid cells in culture

    SciTech Connect

    Isozaki, O.; Kohn, L.D. )

    1987-11-01

    Removal of TSH, insulin, and cortisol from the medium, and decreasing the serum content to 0.2%, abolishes both the proliferate and differentiated state of FRTL-5 rat thyroid cells in culture. In these basal conditions, the individual addition of TSH, insulin, insulin-like growth factor-I (IGF-I), phorbol 12-myristate 13-acetate (TPA), alpha 1-adrenergic agents, or A23187, increase c-myc and/or c-fos proto-oncogene expression. Under the same conditions, only the addition of TSH increased cAMP levels; 8-bromo-cAMP can increase c-myc or c-fos mRNA levels. Pretreatment of cells with phorbol 12,13-dibutyrate, an agent which down regulates the C-kinase, completely inhibits the effect of TPA on proto-oncogene expression but has no affect on the A23187 induced-increase. The sum of these results indicate that at least four separate signal systems independently increase c-myc or c-fos gene expression in FRTL-5 cells cAMP (TSH), C-kinase (TPA), Ca++/phosphoinositide (A23187), and that influenced by insulin/IGF-I. None of the ligands, when individually returned to cells in basal medium (no TSH, insulin, or cortisol and only 0.2% serum), increases cell number; norepinephrine, and A23187 do not increase (3H)thymidine incorporation into DNA under these conditions; and combinations of the ligands can be more than additive in effecting (3H)thymidine incorporation into DNA but are only additive in effecting proto-oncogene expression. Insulin/IGF-I plus TSH or insulin/IGF-I plus norepinephrine can increase both proto-oncogene expression and (3H)thymidine incorporation into DNA to the same extent; however, the former combination can increase cell number whereas the latter cannot. There is therefore no simple correlation between the ability of the above ligands to increase proto-oncogene expression and their ability to increase cell number or induce DNA synthesis.

  13. Tuning of alternative splicing--switch from proto-oncogene to tumor suppressor.

    PubMed

    Shchelkunova, Aleksandra; Ermolinsky, Boris; Boyle, Meghan; Mendez, Ivan; Lehker, Michael; Martirosyan, Karen S; Kazansky, Alexander V

    2013-01-01

    STAT5B, a specific member of the STAT family, is intimately associated with prostate tumor progression. While the full form of STAT5B is thought to promote tumor progression, a naturally occurring truncated isoform acts as a tumor suppressor. We previously demonstrated that truncated STAT5 is generated by insertion of an alternatively spliced exon and results in the introduction of an early termination codon. Present approaches targeting STAT proteins based on inhibition of functional domains of STAT's, such as DNA-binding, cooperative binding (protein-protein interaction), dimerization and phosphorylation will halt the action of the entire gene, both the proto-oncogenic and tumor suppressor functions of Stat5B. In this report we develop a new approach aimed at inhibiting the expression of full-length STAT5B (a proto-oncogene) while simultaneously enhancing the expression of STAT5∆B (a tumor suppressor). We have demonstrated the feasibility of using steric-blocking splice-switching oligonucleotides (SSOs) with a complimentary sequence to the targeted exon-intron boundary to enhance alternative intron/exon retention (up to 10%). The functional effect of the intron/exon proportional tuning was validated by cell proliferation and clonogenic assays. The new scheme applies specific steric-blocking splice-switching oligonucleotides and opens an opportunity for anti-tumor treatment as well as for the alteration of functional abilities of other STAT proteins. PMID:23289016

  14. Tuning of Alternative Splicing - Switch From Proto-Oncogene to Tumor Suppressor

    PubMed Central

    Shchelkunova, Aleksandra; Ermolinsky, Boris; Boyle, Meghan; Mendez, Ivan; Lehker, Michael; Martirosyan, Karen S.; Kazansky, Alexander V.

    2013-01-01

    STAT5B, a specific member of the STAT family, is intimately associated with prostate tumor progression. While the full form of STAT5B is thought to promote tumor progression, a naturally occurring truncated isoform acts as a tumor suppressor. We previously demonstrated that truncated STAT5 is generated by insertion of an alternatively spliced exon and results in the introduction of an early termination codon. Present approaches targeting STAT proteins based on inhibition of functional domains of STAT's, such as DNA-binding, cooperative binding (protein-protein interaction), dimerization and phosphorylation will halt the action of the entire gene, both the proto-oncogenic and tumor suppressor functions of Stat5B. In this report we develop a new approach aimed at inhibiting the expression of full-length STAT5B (a proto-oncogene) while simultaneously enhancing the expression of STAT5∆B (a tumor suppressor). We have demonstrated the feasibility of using steric-blocking splice-switching oligonucleotides (SSOs) with a complimentary sequence to the targeted exon-intron boundary to enhance alternative intron/exon retention (up to 10%). The functional effect of the intron/exon proportional tuning was validated by cell proliferation and clonogenic assays. The new scheme applies specific steric-blocking splice-switching oligonucleotides and opens an opportunity for anti-tumor treatment as well as for the alteration of functional abilities of other STAT proteins. PMID:23289016

  15. Tribbles breaking bad: TRIB2 suppresses FOXO and acts as an oncogenic protein in melanoma.

    PubMed

    Link, Wolfgang

    2015-10-01

    TRIB2 (tribbles homolog 2) encodes one of three members of the tribbles family in mammals. These members share a Trb (tribbles) domain, which is homologous to protein serine-threonine kinases, but lack the active site lysine. The tribbles proteins interact and modulate the activity of signal transduction pathways in a number of physiological and pathological processes. TRIB2 has been identified as an oncogene that inactivates the transcription factor CCAAT/enhancer-binding protein α (C/EBPα) and causes acute myelogenous leukaemia (AML). Recent research provided compelling evidence that TRIB2 can also act as oncogenic driver in solid tumours, such as lung and liver cancer. In particular, our recent work demonstrated that TRIB2 is dramatically overexpressed in malignant melanomas compared with normal skin and promotes the malignant phenotype of melanoma cells via the down-regulation of FOXO (forkhead box protein O) tumour suppressor activity in vitro and in vivo. TRIB2 was found to be expressed in normal skin, but its expression consistently increased in benign nevi, melanoma and was highest in samples from patients with malignant melanoma. The observation that TRIB2 strongly correlates with the progression of melanocyte-derived malignancies suggests TRIB2 as a meaningful biomarker to both diagnose and stage melanoma. In addition, interfering with TRIB2 activity might be a therapeutic strategy for the treatment of several different tumour types. PMID:26517928

  16. Inhibition of the Pim1 Oncogene Results in Diminished Visual Function

    PubMed Central

    Yin, Jun; Shine, Lisa; Raycroft, Francis; Deeti, Sudhakar; Reynolds, Alison; Ackerman, Kristin M.; Glaviano, Antonino; O'Farrell, Sean; O'Leary, Olivia; Kilty, Claire; Kennedy, Ciaran; McLoughlin, Sarah; Rice, Megan; Russell, Eileen; Higgins, Desmond G.; Hyde, David R.; Kennedy, Breandan N.

    2012-01-01

    Our objective was to profile genetic pathways whose differential expression correlates with maturation of visual function in zebrafish. Bioinformatic analysis of transcriptomic data revealed Jak-Stat signalling as the pathway most enriched in the eye, as visual function develops. Real-time PCR, western blotting, immunohistochemistry and in situ hybridization data confirm that multiple Jak-Stat pathway genes are up-regulated in the zebrafish eye between 3–5 days post-fertilisation, times associated with significant maturation of vision. One of the most up-regulated Jak-Stat genes is the proto-oncogene Pim1 kinase, previously associated with haematological malignancies and cancer. Loss of function experiments using Pim1 morpholinos or Pim1 inhibitors result in significant diminishment of visual behaviour and function. In summary, we have identified that enhanced expression of Jak-Stat pathway genes correlates with maturation of visual function and that the Pim1 oncogene is required for normal visual function. PMID:23300608

  17. PET imaging of epidermal growth factor receptor expression in tumours using 89Zr-labelled ZEGFR:2377 affibody molecules

    PubMed Central

    GAROUSI, JAVAD; ANDERSSON, KEN G.; MITRAN, BOGDAN; PICHL, MARIE-LOUISE; STÅHL, STEFAN; ORLOVA, ANNA; LÖFBLOM, JOHN; TOLMACHEV, VLADIMIR

    2016-01-01

    Epidermal growth factor receptor (EGFR) is a transmembrane tyrosine kinase receptor, which is overexpressed in many types of cancer. The use of EGFR-targeting monoclonal antibodies and tyrosine-kinase inhibitors improves significantly survival of patients with colorectal, non-small cell lung cancer and head and neck squamous cell carcinoma. Detection of EGFR overexpression provides important prognostic and predictive information influencing management of the patients. The use of radionuclide molecular imaging would enable non-invasive repeatable determination of EGFR expression in disseminated cancer. Moreover, positron emission tomography (PET) would provide superior sensitivity and quantitation accuracy in EGFR expression imaging. Affibody molecules are a new type of imaging probes, providing high contrast in molecular imaging. In the present study, an EGFR-binding affibody molecule (ZEGFR:2377) was site-specifically conjugated with a deferoxamine (DFO) chelator and labelled under mild conditions (room temperature and neutral pH) with a positron-emitting radionuclide 89Zr. The 89Zr-DFO-ZEGFR:2377 tracer demonstrated specific high affinity (160±60 pM) binding to EGFR-expressing A431 epidermoid carcinoma cell line. In mice bearing A431 xenografts, 89Zr-DFO-ZEGFR:2377 demonstrated specific uptake in tumours and EGFR-expressing tissues. The tracer provided tumour uptake of 2.6±0.5% ID/g and tumour-to-blood ratio of 3.7±0.6 at 24 h after injection. 89Zr-DFO-ZEGFR:2377 provides higher tumour-to-organ ratios than anti-EGFR antibody 89Zr-DFO-cetuximab at 48 h after injection. EGFR-expressing tumours were clearly visualized by microPET using 89Zr-DFO-ZEGFR:2377 at both 3 and 24 h after injection. In conclusion, 89Zr-DFO-ZEGFR:2377 is a potential probe for PET imaging of EGFR-expression in vivo. PMID:26847636

  18. PET imaging of epidermal growth factor receptor expression in tumours using 89Zr-labelled ZEGFR:2377 affibody molecules.

    PubMed

    Garousi, Javad; Andersson, Ken G; Mitran, Bogdan; Pichl, Marie-Louise; Ståhl, Stefan; Orlova, Anna; Löfblom, John; Tolmachev, Vladimir

    2016-04-01

    Epidermal growth factor receptor (EGFR) is a transmembrane tyrosine kinase receptor, which is overexpressed in many types of cancer. The use of EGFR-targeting monoclonal antibodies and tyrosine-kinase inhibitors improves significantly survival of patients with colorectal, non-small cell lung cancer and head and neck squamous cell carcinoma. Detection of EGFR overexpression provides important prognostic and predictive information influencing management of the patients. The use of radionuclide molecular imaging would enable non-invasive repeatable determination of EGFR expression in disseminated cancer. Moreover, positron emission tomography (PET) would provide superior sensitivity and quantitation accuracy in EGFR expression imaging. Affibody molecules are a new type of imaging probes, providing high contrast in molecular imaging. In the present study, an EGFR-binding affibody molecule (ZEGFR:2377) was site-specifically conjugated with a deferoxamine (DFO) chelator and labelled under mild conditions (room temperature and neutral pH) with a positron-emitting radionuclide (89)Zr. The (89)Zr-DFO-ZEGFR:2377 tracer demonstrated specific high affinity (160 ± 60 pM) binding to EGFR-expressing A431 epidermoid carcinoma cell line. In mice bearing A431 xenografts, (89)Zr-DFO-ZEGFR:2377 demonstrated specific uptake in tumours and EGFR-expressing tissues. The tracer provided tumour uptake of 2.6 ± 0.5% ID/g and tumour-to-blood ratio of 3.7 ± 0.6 at 24 h after injection. (89)Zr-DFO-ZEGFR:2377 provides higher tumour-to-organ ratios than anti-EGFR antibody (89)Zr-DFO-cetuximab at 48 h after injection. EGFR‑expressing tumours were clearly visualized by microPET using (89)Zr-DFO-ZEGFR:2377 at both 3 and 24 h after injection. In conclusion, 8(9)Zr-DFO-ZEGFR:2377 is a potential probe for PET imaging of EGFR-expression in vivo. PMID:26847636

  19. Development of Cu-64 labeled EGF for In Vivo PET Imaging of EGFR Expression

    SciTech Connect

    Backer, Joseph M.

    2009-07-12

    In this project we proposed to establish feasibility of the development of targeted tracers for radionuclide imaging of epidermal growth factor receptors (EGFR) in cancer patients. The significance and impact of the proposed radiotracers are determined by the crucial role that EGFR plays in many cancers and by the rapid entrance of EGFR-inhibiting drugs into clinic. Clinical experience, however, revealed that only 10-25% of patients that are defined as EGFR-positive by immunohistochemical analysis respond to EGFR-directed therapeutics and there is poor correlation between EGFR immunohistochemistry and treatment. Therefore, for more efficacious use of EGFR-targeting therapeutics, there is a need for information about EGFR activity in patients. We hypothesized that radionuclide imaging of functionally active EGFR will provide such information and would allow for 1) rational patient stratification, 2) rapid monitoring of responses to therapy, and 3) development of personalized treatment regimens. We hypothesized that tracers based epidermal growth factor (EGF), a natural EGFR ligand, as a targeting vector would be particularly advantageous. First, only functionally active and therefore critical for disease progression EGFRs will bind and internalize an EGF-based tracer. Second, continuous internalization of EGF-based tracers by recyclable EGFR would lead to intracellular accumulation of radionuclide and improved signal-to-background ratio. Third, small size of EGF relative to antibodies would facilitate tumor penetration with vastly better non-specific soft tissue and blood clearance rates. Fourth, as a human protein, EGF is not expected to be immunogenic. Finally, at the beginning of this project, we have already engineered and expressed functionally active EGF with an N-terminal Cys-tag for site-specific conjugation of various payloads, including radionuclide chelators. In the Phase I of this project, in collaboration with Dr. Blankenberg’s group at Stanford

  20. Adolescent Self-Esteem and Gender: Exploring Relations to Sexual Harassment, Body Image, Media Influence, and Emotional Expression.

    ERIC Educational Resources Information Center

    Polce-Lynch, Mary; Myers, Barbara J.; Kliewer, Wendy; Kilmartin, Christopher

    2001-01-01

    Evaluated self-reported influences on self-esteem involving the media, sexual harassment, body image, family and peer relationships, and emotional expression for 93 boys and 116 girls in grades 5, 8, and 12. Results generally supported a pattern in which boys and girls were most similar in late childhood and again in late adolescence. Discusses…

  1. Calcium imaging of vomeronasal organ response using slice preparations from transgenic mice expressing G-CaMP2.

    PubMed

    Yu, C Ron

    2013-01-01

    The vomeronasal organ (VNO) in vertebrate animals detects pheromones and interspecies chemical signals. We describe in this chapter a Ca(2+) imaging approach using transgenic mice that express the genetically encoded Ca(2+) sensor G-CaMP2 in VNO tissue. This approach allows us to analyze the complex patterns of the vomeronasal neuron response to large number of chemosensory stimuli. PMID:24014364

  2. Pinpointing retrovirus entry sites in cells expressing alternatively spliced receptor isoforms by single virus imaging

    PubMed Central

    2014-01-01

    Background The majority of viruses enter host cells via endocytosis. Current knowledge of viral entry pathways is largely based upon infectivity measurements following genetic and/or pharmacological interventions that disrupt vesicular trafficking and maturation. Imaging of single virus entry in living cells provides a powerful means to delineate viral trafficking pathways and entry sites under physiological conditions. Results Here, we visualized single avian retrovirus co-trafficking with markers for early (Rab5) and late (Rab7) endosomes, acidification of endosomal lumen and the resulting viral fusion measured by the viral content release into the cytoplasm. Virus-carrying vesicles either merged with the existing Rab5-positive early endosomes or slowly accumulated Rab5. The Rab5 recruitment to virus-carrying endosomes correlated with acidification of their lumen. Viral fusion occurred either in early (Rab5-positive) or intermediate (Rab5- and Rab7-positive) compartments. Interestingly, different isoforms of the cognate receptor directed virus entry from distinct endosomes. In cells expressing the transmembrane receptor, viruses preferentially entered and fused with slowly maturing early endosomes prior to accumulation of Rab7. By comparison, in cells expressing the GPI-anchored receptor, viruses entered both slowly and quickly maturing endosomes and fused with early (Rab5-positive) and intermediate (Rab5- and Rab7-positive) compartments. Conclusions Since the rate of low pH-triggered fusion was independent of the receptor isoform, we concluded that the sites of virus entry are determined by the kinetic competition between endosome maturation and viral fusion. Our findings demonstrate the ability of this retrovirus to enter cells via alternative endocytic pathways and establish infection by releasing its content from distinct endosomal compartments. PMID:24935247

  3. A transgenic red fluorescent protein-expressing nude mouse for color-coded imaging of the tumor microenvironment.

    PubMed

    Yang, Meng; Reynoso, Jose; Bouvet, Michael; Hoffman, Robert M

    2009-02-01

    The tumor microenvironment (TME) is critical for tumor growth and progression. We have previously developed color-coded imaging of the TME using a green fluorescent protein (GFP) transgenic nude mouse as a host. However, most donor sources of cell types appropriate for study in the TME are from mice expressing GFP. Therefore, a nude mouse expressing red fluorescent protein (RFP) would be an appropriate host for transplantation of GFP-expressing stromal cells as well as double-labeled cancer cells expressing GFP in the nucleus and RFP in the cytoplasm, thereby creating a three-color imaging model of the TME. The RFP nude mouse was obtained by crossing non-transgenic nude mice with the transgenic C57/B6 mouse in which the beta-actin promoter drives RFP (DsRed2) expression in essentially all tissues. In crosses between nu/nu RFP male mice and nu/+ RFP female mice, the embryos fluoresced red. Approximately 50% of the offspring of these mice were RFP nude mice. In the RFP nude mouse, the organs all brightly expressed RFP, including the heart, lungs, spleen, pancreas, esophagus, stomach, duodenum, the male and female reproductive systems; brain and spinal cord; and the circulatory system, including the heart, and major arteries and veins. The skinned skeleton highly expressed RFP. The bone marrow and spleen cells were also RFP positive. GFP-expressing human cancer cell lines, including HCT-116-GFP colon cancer and MDA-MB-435-GFP breast cancer were orthotopically transplanted to the transgenic RFP nude mice. These human tumors grew extensively in the transgenic RFP nude mouse. Dual-color fluorescence imaging enabled visualization of human tumor-host interaction. The RFP nude mouse model should greatly expand our knowledge of the TME. PMID:19097136

  4. Multiple endocrine neoplasias type 2B and RET proto-oncogene

    PubMed Central

    2012-01-01

    Multiple Endocrine Neoplasia type 2B (MEN 2B) is an autosomal dominant complex oncologic neurocristopathy including medullary thyroid carcinoma, pheochromocytoma, gastrointestinal disorders, marphanoid face, and mucosal multiple ganglioneuromas. Medullary thyroid carcinoma is the major cause of mortality in MEN 2B syndrome, and it often appears during the first years of life. RET proto-oncogene germline activating mutations are causative for MEN 2B. The 95% of MEN 2B patients are associated with a point mutation in exon 16 (M918/T). A second point mutation at codon 883 has been found in 2%-3% of MEN 2B cases. RET proto-oncogene is also involved in different neoplastic and not neoplastic neurocristopathies. Other RET mutations cause MEN 2A syndrome, familial medullary thyroid carcinoma, or Hirschsprung's disease. RET gene expression is also involved in Neuroblastoma. The main diagnosis standards are the acetylcholinesterase study of rectal mucosa and the molecular analysis of RET. In our protocol the rectal biopsy is, therefore, the first approach. RET mutation detection