bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract LB-322: The senescence-associated Sin3B protein promotes inflammation and pancreatic cancer progression
Authors: Rielland Maite , Cantor David , Graveline Richard , Hadju Cristina , Mara Lisa , Miller George , David Gregory
Published: May 2015

Pancreatic ductal adenocarcinoma (PDAC) is strikingly resistant to conventional therapeutic approaches. We have previously demonstrated that the histone deacetylase associated SIN3B protein is essential for oncogene-induced senescence in cultured cells. Here we report that SIN3B is required for activated KRAS-induced senescence in vivo using a mouse model of pancreatic cancer. Surprisingly, impaired senescence caused by genetic inactivation of Sin3B is associated with delayed PDAC progression, correlating with an impaired inflammatory response. At the molecular level, SIN3B mediates KRAS-induced Il-1a production in a cell autonomous and senescence dependent manner. These results indicate for the first time that senescence-associated inflammation positively correlates with PDAC progression and identify SIN3B as a therapeutic target to inhibit inflammation-driven tumorigenesis.

Citation Format: Maite Rielland, David Cantor, Richard Graveline, Cristina Hadju, Lisa Mara, George Miller, Gregory David. The senescence-associated Sin3B protein promotes inflammation and pancreatic cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-322. doi:10.1158/1538-7445.AM2014-LB-322
Abstract LB-326: Structural basis of lipid-binding and regulation in PI3Kα
Authors: Miller Michelle S. , Schmidt-Kittler Oleg , Bolduc David M. , Brower Evan T. , Chaves-Moreira Daniele , Allaire Marc , Kinzler Kenneth W. , Jennings Ian G. , Thompson Philip E. , Cole Philip A. , Amzel L. Mario , Vogelstein Bert , Gabelli Sandra B.
Published: May 2015

PI3Kα is a well-established target for the development of novel cancer therapeutics. Currently, all inhibitors in clinical trials target the highly conserved ATP-binding site, which has made the development of PI3K selective inhibitors difficult, with many inhibitors displaying cross-reactivity against protein kinases.

We have used X-ray crystallography and fluorescence quenching studies to characterize the lipid-binding site of PI3Kα and the structural basis of regulation by nSH2. A newly determined p110α/niSH2 crystal structure is the first to reveal the nSH2 domain in complex with wild-type p110α, allowing investigation of the mechanisms of nSH2 regulation. Key interactions between the nSH2 domain and the activation loop suggest a mechanism by which the kinase domain is kept in an inactive conformation until activation by phosphopeptide binding. Key differences in nSH2 domain binding to p110α were identified between the wild-type and oncogenic mutant, p110αH1047R. Increased buried surface area and two unique salt-bridges are suggestive of tighter regulatory control in the wild-type PI3Kα compared with the oncogenic mutant. A second structure reveals the details of PI3K binding to a lipid substrate mimetic, diC4-PIP2. Unexpectedly, we found an additional lipid-binding site, and this striking observation was confirmed by fluorescence quenching experiments. The identification of multiple lipid binding sites provides additional targets that may enable more selective inhibition among the various isoforms, or even between mutant and wild-type forms of PI3Kα.

Citation Format: Michelle S. Miller, Oleg Schmidt-Kittler, David M. Bolduc, Evan T. Brower, Daniele Chaves-Moreira, Marc Allaire, Kenneth W. Kinzler, Ian G. Jennings, Philip E. Thompson, Philip A. Cole, L. Mario Amzel, Bert Vogelstein, Sandra B. Gabelli. Structural basis of lipid-binding and regulation in PI3Kα. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-326. doi:10.1158/1538-7445.AM2014-LB-326
Abstract LB-330: Four individually druggable Met hotspots mediate HGF-driven tumor progression
Authors: Hultberg Anna , Basilico Cristina , Blanchetot Cristophe , Jonge Natalie D. , Hanssens Valérie , Boeck Gitte D. , Mira Alessia , Cazzanti Manuela , Morello Virginia , Dreier Torsten , Saunders Michael , Haard Hans D. , Michieli Paolo
Published: May 2015

Activation of Met by HGF plays a key role in tumor progression. Using a newly developed llama platform generating human-like immunoglobulins, we selected 68 different antibodies that competed with HGF for binding to Met. HGF-competing antibodies recognized four hotspots localized in different domains of Met. One hotspot coincides with the previously identified HGF β-chain binding site on blades 2-3 of the SEMA β-propeller. Two hotspots lie within SEMA blade 5 and IPT domains 2-3, respectively, where the HGF α-chain is thought to bind. The last hotspot identifies a new region across the PSI-IPT 1 domains. Individual or combined targeting of these hotspots effectively interrupted HGF-Met signaling in multiple biochemical and biological assays. Selected antibodies directed against SEMA blades 2-3 and the PSI-IPT 1 region inhibited brain invasion and prolonged survival in a glioblastoma multiforme model, prevented metastatic disease following neo-adjuvant therapy in a triple-negative mammary carcinoma model, and suppressed cancer cell dissemination to the liver in a KRAS-mutant metastatic colorectal cancer model. These results identify multiple regions of Met responsible for HGF-mediated tumor progression, unraveling the complexity of HGF-Met interaction and providing new molecular tools for targeting Met activity in cancer.

Citation Format: Anna Hultberg, Cristina Basilico, Cristophe Blanchetot, Natalie De Jonge, Valérie Hanssens, Gitte De Boeck, Alessia Mira, Manuela Cazzanti, Virginia Morello, Torsten Dreier, Michael Saunders, Hans De Haard, Paolo Michieli. Four individually druggable Met hotspots mediate HGF-driven tumor progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-330. doi:10.1158/1538-7445.AM2014-LB-330
Abstract LB-34: 3-Dimensional air-liquid interface organoid culture of primary human tumor biopsies
Authors: Neal James T. , Cantrell Michael , Rack Paul , Kuo Calvin J.
Published: May 2015

The application of primary organoid cultures 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. We have previously described a single air-liquid interface culture method that can be used to engineer oncogenic mutations into primary epithelial/mesenchymal organoids from mouse colon, stomach, and pancreas (Ootani et al., Nature Medicine 15(6):701-6 (2009); Li et al., in press). We have recently modified this methodology to allow the robust long-term 3-dimensional culture of a wide variety of primary neoplasms from human biopsy samples. These primary tumor organoids accurately recapitulate the architecture and histopathology of the tumors from which they were derived, including epithelial and mesenchymal compartments. These tumor organoids can be further passaged into 96-well format to enable a variety of genetic and pharmacologic manipulations with the overall goal of enabling mechanistic studies of tumorigenesis and obtaining clinically actionable information regarding chemosensitivity and resistance to therapy.

Citation Format: James T. Neal, Michael Cantrell, Paul Rack, Calvin J. Kuo. 3-Dimensional air-liquid interface organoid culture of primary human tumor biopsies. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-34. doi:10.1158/1538-7445.AM2014-LB-34
Abstract LB-40: Inducible in vivo silencing of Brd4 identifies potential toxicities of sustained BET protein inhibition
Authors: Bolden Jessica , Tasdemir Nilgun , Dow Lukas , Es Johan H. van , Wilkinson John E. , Zhao Zhen , Clevers Hans , Lowe Scott W.
Published: May 2015

BET family proteins are novel therapeutic targets for cancer and inflammation and represent the first epigenetic readers against which small-molecule inhibitors have been developed. First generation BET inhibitors have shown some therapeutic efficacy in pre-clinical models, however the consequences of disrupting BET protein function in normal tissues are largely unknown. Using an inducible and reversible transgenic RNAi mouse model targeting the BET family protein Brd4, we show that acute suppression of Brd4 in adult animals has immediate and dramatic effects in multiple tissues. Brd4-depleted mice displayed epidermal hyperplasia, alopecia, and decreased cellular diversity and stem cell depletion in the small intestine. To test whether the loss of intestinal stem cells induced by Brd4 suppression would have consequences in combination with agents that induce acute gut toxicity, we subjected mice to a sub-lethal irradiation. Whereas control mice showed intestinal recovery 4-5 days following irradiation, Brd4-depleted mice showed dramatic and continued crypt-villus atrophy, implying that potent BET protein inhibition in combination with cytotoxic chemotherapy may show significantly enhanced toxicity in the clinic. These findings provide important insight into Brd4 function in normal tissues and importantly, predict several potential outcomes associated with potent and sustained inhibition of BET protein function.

Citation Format: Jessica Bolden, Nilgun Tasdemir, Lukas Dow, Johan H. van Es, John E. Wilkinson, Zhen Zhao, Hans Clevers, Scott W. Lowe. Inducible in vivo silencing of Brd4 identifies potential toxicities of sustained BET protein inhibition. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-40. doi:10.1158/1538-7445.AM2014-LB-40
Abstract LB-44: Persistent androgen ablation promotes enhanced neuroendocrine features in Pten-deficient prostate cancer
Authors: Mulholland David J.
Published: May 2015

Background: The development of potent androgens has increased survival of CRPC patients. Clinical data also suggests that the number of neuroendocrine (NE) CaPs frequently found in soft tissue metastasis has steadily increased over recent years and may be correlated with the use of potent androgen receptor (AR) inhibitors such as Enzalutamide or Aberaterone Acetate. These data suggest that NE cells may escape hormonal castration and that active inhibition of the AR may promote aggressive disease. Approach. To investigate this hypothesis, we have taken both genetic and pharmacological approaches to oppose AR signaling in Pten deficient CaP. Surgical castration of Pten-null mutant mice (Pb-Cre+;PtenL/L) led to both an increased EMT signature but also enhanced content of NE cells that were Synaptophysin+ and Pten-low/null. Chronic Enzalutamide treatment (4 wks., 10 mg/kg/d) promoted an increased percentage Synaptophysin+ cells, an observation that was potentiated when coinciding with surgical castration. Interestingly, the highest potentiation of NE cells was in the proximal anterior region of Pten-null mice. Consistent with our previous results, epithelial deletion of AR signaling in Pten-null epithelium (Pb-Cre+;PtenL/L;ArL/Y) did not prevent CaP progression under surgical castrate conditions. Moreover, compared to Pten-null mutants, Pten-null;Ar-null mutants showed an enhanced stem cell signature (p63+ cells, LSChigh subpopulation) but also marked increase content of NE cells upon both surgical castration and chronic Enzalutamide treatment (4-8 wks., 10 mg/kg/d). To consider whether total systemic ablation of the AR signaling axis could impact progression, resected Pten-null explants (“chuncks”) were isolated from Pb-Cre+;PtenL/L mutants and transplanted to the kidney capsule of Tfm;Rag1-null mice, immune incompetent hosts that are genetically null for Ar. Consistent with our hypothesis, Tfm;Rag1-null mutants showed more NE cells in explants than those transplanted to control AR intact control (Rag1-null) mice. Conclusion: Chronic blockade of the AR signaling axis results in a poorly differentiated phenotype characterized by increased stemness and enhanced NE phenotype.

Citation Format: David J. Mulholland. Persistent androgen ablation promotes enhanced neuroendocrine features in Pten-deficient prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-44. doi:10.1158/1538-7445.AM2014-LB-44
Abstract LB-45: VEGFR3 signaling facilitates mammary tumor cells to resist chemotherapy
Authors: Sharma Bhawna , Varney Michelle L. , Singh Rakesh K.
Published: May 2015

Breast cancer is a deadly disease which accounts for more than 40,000 deaths each year in United States. With the availability of various treatment options, side effects and resistance associated with them is also becoming a major concern for the treatment of breast cancer. How breast cancer cells evade chemotherapy still remains a vast area for research. In this study, we proposed that mammary tumor cells resist cell death via VEGFR3 signaling pathway. To test our hypothesis, we used doxorubicin and paclitaxel resistant murine mammary tumor cells (Cl66-Dox and Cl66-Pac) which have been developed in our laboratory. When implanted in female Balb/c mice they form more aggressive tumor than the parent cells (Cl66) from which they have been derived. We observed that these resistant cells have higher expression of VEGFR3 receptor than parent cells at mRNA level. Treatment with VEGFR3 kinase inhibitor, MAZ51, Cl-66Dox and Cl66-Pac cells showed 50% growth inhibition at 10uM dose, suggesting VEGFR3 receptor activation as one of the mechanism for resistance. Moreover, when implanted in mice, tumor formed by resistant cells responded well to MAZ51 treatment with slower tumor growth in comparison to control group consistent with our in vitro data. However, when we treated mice bearing resistant tumors first with chemotherapeutic drug, docetaxel followed by MAZ51, we observed a significant decrease in tumor growth and tumor weight, examined upon endpoint analysis. These results suggest that combination treatment is more effective in inhibiting drug-resistant tumors growth in comparison to drug and inhibitor alone as determined by reduced tumor volume and weight in combination treatment group. Overall, our in vivo data showed that blocking VEGFR3 signaling pathway makes drug-resistant mammary tumor cells susceptible to chemotherapy suggesting the use of inhibitors for this signaling pathway to be beneficial for drug-resistant mammary tumors.

Citation Format: Bhawna Sharma, Michelle L. Varney, Rakesh K. Singh. VEGFR3 signaling facilitates mammary tumor cells to resist chemotherapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-45. doi:10.1158/1538-7445.AM2014-LB-45
Abstract LB-47: Specific and potent silencing of K-Ras by asymmetric RNA technology reveals addiction of gastric cancer stem cells
Authors: Sun Xiangao , Li Youzhi , Umehara Hiroki , Oishi Jun , Jesuraj Nithya , Barbulovic Jelena , Dai Xiaoshu , Gada Keyur , Li Chiang
Published: May 2015

The protein K-Ras is a molecular switch that under normal conditions regulates cell growth and cell division. Mutations in this protein lead to the formation of tumors through continuous cell growth. About 30% of human cancers have a mutated Ras protein that is constitutively bound to GTP due to decreased GTPase activity and insensitivity to GAP action. Ras is also an important factor in many cancers in which it is not mutated but rather functionally activated through inappropriate activity of other signal transduction elements. Mutated K-Ras proteins are found in a large proportion of all tumour cells. K-Ras protein occupies a central position of interest. The identification of oncogenically mutated K-Ras in many human cancers led to major efforts to target this constitutively activated protein as a rational and selective treatment. Despite decades of active agent research, significant challenges still remain to develop therapeutic inhibitors of K-Ras.

To elucidate the function of K-Ras in the cancer development and maintenance, we developed asymmetric interfering RNAs (aiRNAs) which are able to silence target genes with high potency leading to long-lasting knockdown, and reducing off-target effects, and investigated the dependency of K-ras on cell survival in several types of human cancer cell lines. Much to our surprise, we found K-Ras plays a more significant role for gastric cancer maintenance compared to other types of cancer. Here we report aiRNA-induced silencing of K-Ras inhibited the cell proliferation of gastric cancer cells and the ability of gastric cancer cells to form colonies compared to other cancer types. Accumulating evidence has revealed that cancer stem cells (CSCs) are highly associated with prognosis, metastasis, and recurrence. To investigate the effect of K-Ras on CSCs, we tested the K-Ras gene silencing effects on an in vitro CSC culturing system. As a result, K-Ras inhibition decreased the colonies derived from gastric CSCs and altered the gene expression patterns of several genes involved in “stemness” compared to other cancer types. The results of these studies suggest that gastric cancer and gastric CSCs are affected by the K-Ras oncogene and that Kras aiRNAs are promising therapeutic candidates for the treatment of gastric cancer.

Citation Format: Xiangao Sun, Youzhi Li, Hiroki Umehara, Jun Oishi, Nithya Jesuraj, Jelena Barbulovic, Xiaoshu Dai, Keyur Gada, Chiang Li. Specific and potent silencing of K-Ras by asymmetric RNA technology reveals addiction of gastric cancer stem cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-47. doi:10.1158/1538-7445.AM2014-LB-47
Abstract LB-52: CBX7 inhibits breast cancer stem cell activity and tumorigenicity through DKK-1-mediated suppression of the Wnt/β-catenin pathway
Authors: Kim Hey-Yon , Park Ji-Hye , Won Hee-Young , Lee Jeong-Yeon , Kong Gu
Published: May 2015

Chromoprotein homologue 7 (CBX7), a member of the polycomb repressive complex (PRC)-1, has been involved in several biological processes including stem cell regulation and cancer development, but its roles in cancer stem cells (CSCs) and breast tumorigenicity remain unknown. Here, we demonstrated that CBX7 negatively regulates breast CSCs. CD44+/CD24-/ESA+ cells, a population of breast CSCs, showed diminished CBX7 expression compared with non-CSCs. Furthermore, small hairpin RNA (shRNA)-mediated CBX7 knockdown in breast epithelial and cancer cell lines increased breast CSC population and reinforced in vitro self-renewal and in vivo tumor initiating ability. Similarly, CBX7 overexpression showed reduced stemness of breast CSCs. We also found that CBX7 inhibits the Wnt/β-catenin/TCF pathway by enhancing the expression of Dickkopf-1 (DKK-1), a Wnt antagonist, to impede self-renewal of breast CSCs. Particularly, CBX7 increased DKK-1 transcription by cooperating with p300 acetyltransferase and subsequently enhancing the histone acetylation of DKK-1 promoter. Furthermore, pharmacological inhibition of DKK-1 in CBX7 overexpressing cells showed recovery of the Wnt signaling and consequent rescue of breast CSC activity. Taken together, these findings indicated that CBX7-mediated epigenetic induction of DKK-1 is crucial for inhibition of breast CSC activity, suggesting that CBX7 could be a potential tumor suppressor as a novel negative regulator of CSCs in human breast cancer.

Citation Format: Hey-Yon Kim, Ji-Hye Park, Hee-Young Won, Jeong-Yeon Lee, Gu Kong. CBX7 inhibits breast cancer stem cell activity and tumorigenicity through DKK-1-mediated suppression of the Wnt/β-catenin pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-52. doi:10.1158/1538-7445.AM2014-LB-52
Abstract LB-54: HOXB7 regulates breast cancer stem cells via induction of TGFβ2
Authors: Liu Shou , Johnson Sarah , Chen Hexin
Published: May 2015

HOX genes encode a family of homeodomain-containing transcription factors involved in the determination of cell fate and identity during embryonic development. HOXB7 is overexpressed in more than 70% of breast cancers. Overexpression of HOXB7 in breast epithelial cells and induces epithelial-mesenchymal transition. In this study, we found that overexpression of the HOXB7 gene in breast cell lines increases cancer stem cell populations and promotes cell invasion, in contrast, knockdown of HOXB7 expression decreases the proportions of cancer stem cells. Colonogenic assay and xenograft transplantation experiments further confirmed that overexpression of HOXB7 in breast cancer cells increases their tumorigenecity both in vitro and in vivo. Mechanistically we found that HOXB7 strongly induces TGFβ2 expression and activates the canonical TGFβ/SMAD signaling pathways, which may be responsible for induction of EMT leading to expansion of cancer stem cell population. Overall, our results suggest that HOXB7 may be a critical factor which regulates the self-renewal and differentiation of breast cancer stem cells and therefore a potential therapeutic target in breast cancer.

Citation Format: Shou Liu, Sarah Johnson, Hexin Chen. HOXB7 regulates breast cancer stem cells via induction of TGFβ2. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-54. doi:10.1158/1538-7445.AM2014-LB-54
Abstract LB-55: UTX epigenetically regulates epithelial-mesenchymal transition-associated breast cancer stem cell properties independently of EZH2
Authors: Choi Hee-Joo , Park Ji-Hye , Park Mi-Kyung , Lee Chang-Hoon , Lee Jeong-Yeon , Kong Gu
Published: May 2015

UTX is a histone H3 lysine 27 (H3K27) demethylase that can antagonize a histone methyltransferase EZH2. Although H3K27 trimethylation by EZH2 has been involved in cancer-related biological processes including tumorigenesis, invasion, and stemness, the role of UTX in cancer progression has not been identified yet. In this study, we demonstrated an EZH2-independent tumor suppressive function of UTX in breast cancer. We found that loss of UTX enhanced self-renewal and tumorigenicity of breast cancer stem cells (CSCs) and also promoted CSC-associated epithelial-mesenchymal transition (EMT) through upregulating EMT-transcription factors (EMT-TFs), Snail, ZEB1 and ZEB2. Moreover, UTX positively regulated E-cadherin, a key marker of EMT, via these EMT-TFs. Notably, we identified that UTX facilitated epigenetic silencing of EMT-TFs by cooperating with c-Myc and various epigenetic modulators independently of EZH2. In the promoter regions of EMT-TFs, the levels of histone H3K4 methylation and H3 acetylation were inhibited by UTX. We also found that UTX formed a transcriptional repressive complex with LSD1, HDAC1, and DNMT1 for recruiting them to the promoters of EMT-TFs, while loss of UTX led to recruitment of c-Myc and p300 transcriptional active complex to these promoters. These effects were dependent on c-Myc, as UTX knockdown-induced epigenetic alterations of EMT-TFs and breast cancer stemness were reversed by siRNA-mediated c-Myc repression. Taken together, these data indicated that UTX negatively regulates EMT-induced breast CSCs by epigenetically repressing EMT-TFs independently of H3K27 methylation, suggesting a novel tumor suppressive role of UTX in breast cancer.

Citation Format: Hee-Joo Choi, Ji-Hye Park, Mi-Kyung Park, Chang-Hoon Lee, Jeong-Yeon Lee, Gu Kong. UTX epigenetically regulates epithelial-mesenchymal transition-associated breast cancer stem cell properties independently of EZH2. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-55. doi:10.1158/1538-7445.AM2014-LB-55
Abstract LB-66: NVP-BEZ235 and bevacizumab as therapy for glioblastoma
Authors: Netland Inger Anne , Førde Hilde , Sleire Linda , Skeie Bente S. , Enger Per Øyvind , Goplen Dorota
Published: May 2015

Glioblastoma (GBM) is the most common and aggressive primary brain tumour. Its dismal prognosis despite multimodal treatment comprising surgery, radiotherapy and chemotherapy, calls for new therapeutic strategies.

GBM is a highly vascularized cancer, with elevated expression levels of vascular endothelial growth factor (VEGF), a key mediator of angiogenesis. Bevacizumab is a humanized anti-VEGF monoclonal antibody in clinical use as an angiogenesis inhibitor. Although bevacizumab is approved for treatment of GBM, several studies now show that it does not prolong survival. In addition, it has been shown that bevacizumab treatment leads to increased invasion and upregulation of gene transcripts involved in phosphoinositol-3-kinase (PI3K) - pathways. NVP-BEZ235 is a dual PI3K/mTOR kinase inhibitor. Combining NVP-BEZ235 with bevacizumab may therefore target both the angiogenic and invasive phenotype of GBMs.In vitro, the U-87 glioma cell line showed a dose-dependent reduction in cell proliferation to doses of NVP-BEZ235 from 0 to 250nM, as shown by direct cell count, BrdU incorporation and MTS assay. Furthermore, this was accompanied by a dose-dependent reduction of phosphorylated Akt at Serine 473, while the total levels of Akt remained unchanged. An IC50 of 11nM NVP-BEZ235 for the tumour material implanted, was determined in vitro by MTS assay. In order to validate combination treatment of BEZ235 and Avastin in vivo, we utilized a clinically relevant patient-based GBM xenograft model in NOD/SCID mice that mimics both the invasive and angiogenic growth pattern of human GBMs. In total 40 mice were allocated to four groups that received combination treatment with Avastin/BEZ235, monotreatment with Avastin or BEZ235 and untreated controls. Tumor engraftment and growth was confirmed and monitored longitudinally with MRI. The study is ongoing, survival data including histopathological analysis will be presented.

Citation Format: Inger Anne Netland, Hilde Førde, Linda Sleire, Bente S. Skeie, Per Øyvind Enger, Dorota Goplen. NVP-BEZ235 and bevacizumab as therapy for glioblastoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-66. doi:10.1158/1538-7445.AM2014-LB-66
Abstract LB-68: MYCN-induced TRPM7 expression and channel activity occurs through a mechanism that involves ornithine decarboxylase
Authors: Koomoa Dana-Lynn T. , Lange Ingo
Published: May 2015

Neuroblastoma (NB) is an extra-cranial solid cancer in children. MYCN is a transcription factor that regulates the expression of genes that are involved in cell proliferation, growth, cell migration, etc. MYCN gene amplification is a prognostic indicator of poor outcome in NB. Recent studies have shown that intracellular free calcium regulates multiple steps involved in cell migration. Although significant advances have been made in understanding the role of calcium during migration, little has been achieved towards understanding its impact on the malignant progression of NB. Our results showed that MYCN gene amplification or over-expression of MYCN in NB cells increased TRPM7 expression, increased peak TRPM7 current, induced constitutive TRPM7 channel activity, enhanced calcium signaling, and promoted cell migration, implicating TRPM7 as a potential regulator of calcium signals that promote NB cell migration and invasion. Recent results suggest that MYCN may regulate TRPM7 expression through a process that involves ornithine decarboxylase (ODC), a rate limiting enzyme in the biosynthesis of polyamines, and a direct transcriptional target of MYCN. Treatment of NB cells with α-difluoromethylornithine (DFMO), an irreversible inhibitor of ODC, or transfection of ODC specific siRNA effectively decreased TRPM7 expression. This effect was attenuated by supplementing the culture media with spermidine, confirming that ODC may function downstream of MYCN to regulate TRPM7 expression. Interestingly, western blot analysis of NB cell lysates showed that MYCN induced the cleavage of the TRPM7 kinase domain, a post-translational modification that has been shown to increase TRPM7 currents and induce constitutive TRPM7 channel activity. Additionally, DFMO inhibited TRPM7 cleavage, an effect that was reversed by spermidine supplementation. Taken together, these results suggest that MYCN may regulate TRPM7 expression and channel activity through a mechanism that involves ODC-mediated polyamine biosynthesis.

Citation Format: Dana-Lynn T. Koomoa, Ingo Lange. MYCN-induced TRPM7 expression and channel activity occurs through a mechanism that involves ornithine decarboxylase. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-68. doi:10.1158/1538-7445.AM2014-LB-68
Abstract LB-69: Rab11 mediated focal adhesion turnover in sarcoma cell migration
Authors: Kao Ling-Yi , Wu Yi-Che , Chao Wei-Ting
Published: May 2015

Cell migration is central for many physiological and pathological processes, including embryogenesis, tissue repair, inflammatory responses, and cancer metastasis. Focal adhesion (FA) turnover has been demonstrated to play important role in controlling cell migration. Recently, FA disassembly was shown to be regulated by endocytosis during cell migration. Rabs are GTPase proteins that participate in various cellular vesicle transports; however, whether Rab proteins play the role in mediating vesicle transport for FA formation is not clear. This study is examining whether Rab11 mediated endosome recycling regulates FA formation during cell migration. Rab11 Wild-type, dominant negative or shRNA were transfected into human HT1080 fibrosarcoma cells, the cell motility was determined by trans-well assay and random migration assay, the localization of Rab11 and focal adhesion molecules were monitored by confocal microscopy. Results showed the Rab11 dominant negative form or shRNA transfection inhibited chemoattractive cell migration, however, only the Rab11 shRNA transfection inhibited random cell migration. Rab11 was also found colocalized with recycled β1 integrin and focal adhesion kinase and affected focal adhesion formation. Taken together, the results suggested Rab11 affected cell migration by regulating FAK dynamics and the activity of Rab11 is required for external stimulation. The interaction of Rab11 with associated focal adhesion molecules will be further investigated.

Citation Format: Ling-Yi Kao, Yi-Che Wu, Wei-Ting Chao. Rab11 mediated focal adhesion turnover in sarcoma cell migration. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-69. doi:10.1158/1538-7445.AM2014-LB-69
Abstract LB-79: PRC2 loss amplifies Ras-driven transcription and sensitizes cancers to bromodomain inhibitor-based combination therapies
Authors: Raedt Thomas D. , Beert Eline , Pasmant Eric , Luscan Armelle , Brems Hilde , Ortonne Nicolas , Helin Kristian , Mautner Victor , Kehrer-Sawatski Hildegard , Clapp Wade , Bradner James , Upadhyaya Meena , Vidaud Michel , Legius Eric , Cichowski Karen
Published: May 2015

The polycomb repressive complex 2 (PRC2) exerts oncogenic effects in many tumor types. However, loss-of-function mutations in PRC2 components occur in a subset of hematopoietic malignancies, suggesting that this complex plays a dichotomous and poorly understood role in cancer. Our genomic, cellular, and mouse modeling data demonstrates that the polycomb group gene SUZ12 functions as tumor suppressor in PNS tumors, glioblastomas, and melanomas by cooperating with mutations in NF1. NF1 encodes a Ras GTPase-activating protein and its loss drives cancer by activating Ras. We show that SUZ12-loss potentiates the effects of NF1 mutations by amplifying Ras-driven transcription. Importantly however, SUZ12-inactivation also triggers an epigenetic switch that sensitizes these cancers to bromodomain inhibitors. Collectively, these studies not only reveal an unexpected connection between the PRC2 complex, NF1, and Ras, but also identify a promising epigenetic-based therapeutic strategy that may be exploited for a variety of cancers.

Citation Format: Thomas De Raedt, Eline Beert, Eric Pasmant, Armelle Luscan, Hilde Brems, Nicolas Ortonne, Kristian Helin, Victor Mautner, Hildegard Kehrer-Sawatski, Wade Clapp, James Bradner, Meena Upadhyaya, Michel Vidaud, Eric Legius, Karen Cichowski. PRC2 loss amplifies Ras-driven transcription and sensitizes cancers to bromodomain inhibitor-based combination therapies. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-79. doi:10.1158/1538-7445.AM2014-LB-79
Abstract LB-84: Mutant p53 enhances the metastatic potential of ovarian cancer cells
Authors: Ahn Ji-Hye , Lee Jong-Gyu , Choi Jung-Hye
Published: May 2015

Missense-mutations of TP53 resulting in accumulation of mutant proteins are very frequent in human cancer including ovarian cancer. Aggressive ovarian cancer is characterized by a peritoneal metastasis which involves the adhesion, migration, and invasion of tumor cells to the peritoneal cavity. In the present study, we investigated the effect of p53 GOF mutation (p53R248) on the metastatic potential of human ovarian cancer cells. We found that forced overexpression of p53R248 significantly increased adhesion to mesothelial cell Met5A, migration, and invasion in p53-null SKOV-3 cells. Conversely, knockdown of mutant p53 using siRNA significantly compromised p53R248-induced cell adhesion, migration, and invasion. Microarray analysis revealed that several metastasis-related genes including integrins and S1PR1(Sphingosine-1-phosphate receptor 1), and certain signaling pathway including PI3K/AKT were markedly up-regulated in p53R248 transfectant SKOV-3R248 cells. Inhibition of PI3K/Akt signaling using specific inhibitor significantly attenuated p53R248-mediated mesothelial cell adhesion, migration, and invasion. These finding show for the first time that mutant p53R248 may exert gain-of-function activity to regulate metastasis-related gene expression and the PI3K/Akt singling pathway in ovarian cancer.

Citation Format: Ji-Hye Ahn, Jong-Gyu Lee, Jung-Hye Choi. Mutant p53 enhances the metastatic potential of ovarian cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-84. doi:10.1158/1538-7445.AM2014-LB-84
Abstract LB-85: Damage-inducible intragenic demethylation activates transcription of an alternative intronic promoter in TP53-wildtype but not mutant human cells and tumors
Authors: Blackburn James , Ng Robert , Roden Daniel , Wu Jianmin , Epstein Richard J.
Published: May 2015

Unlike the well-known effects of promoter methylation, the functional significance of altered intragenic (gene body) CpG methylation remains unclear, particularly in genes such as the TP53 tumor suppressor which lack a 5′ CpG island. Here we show that CpG-specific demethylation of TP53 exon 5 in normal human and mouse cells is inducible by X-irradiation; this is accompanied by co-expression of a 5′-truncated TP53 isoform, raising the possibility that demethylation permits transcription from the known alternative promoter (P2) in intron 4. Consistent with this, in vivo expression of fully pre-methylated TP53 exonic constructs is associated with reduced basal and damage-inducible expression of both full-length and truncated TP53. In contrast, virally TP53-reconstituted Caco-2 and PC-3 human cancer cell lines respond to damage by upregulating expression of full-length TP53 and reducing expression of the truncated RNA isoform; CDKN1A expression varies directly with P2 isoform expression but inversely with the P1 transcript in these cells, suggesting that the isoform mediates downstream transactivation. 450K methylation analysis of adjacent normal and TP53-mutated human pancreatic cancer tissues suggests a mutation-specific decrease of intragenic TP53 methylation in intron 1 near P1 (p < 0.02 ) and a reciprocal increase in intron 4 near P2 (p < 0.01). Archival TP53 database analysis indicates that CpG-specific mutation frequency in human tumors is inversely associated with damage-inducible demethylation at the same sites in vitro (Spearman coefficient -7.1 ), consistent with methylation-dependent downregulation of transcription-coupled repair. We conclude that intragenic alterations of TP53 methylation are both physiologically inducible and functional, and hypothesise that tumor progression is drivable by either (i) intragenic hypermethylation affecting P1 as a field defect in TP53-wt tumor or adjacent normal tissues, or (ii) damage-inducible repression of P2 isoform expression associated with defective intron 4 demethylation, together with overexpression of full-length p53, in TP53-mutant tumors and cells.

Citation Format: James Blackburn, Robert Ng, Daniel Roden, Jianmin Wu, Richard J. Epstein. Damage-inducible intragenic demethylation activates transcription of an alternative intronic promoter in TP53-wildtype but not mutant human cells and tumors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-85. doi:10.1158/1538-7445.AM2014-LB-85
Abstract PL04-02: The breast epithelial hierarchy and its implications for tumor heterogeneity
Authors: Visvader Jane E. , Rios Anne , Fu Naiyang , Nolan Emma , Lindeman Geoffrey
Published: May 2015

Breast cancer is a highly heterogeneous disease at both the molecular and pathological levels. To understand this heterogeneity and ‘cells of origin’ of breast cancer, it is important to dissect the normal mammary epithelial hierarchy. Discrete populations of mouse and human mammary epithelial cells can be isolated on the basis of cell surface marker expression. To date, subpopulations that are highly enriched for mammary stem (MaSC), luminal progenitor and mature luminal cells have been identified. Notably, these are functionally analogous across human and mouse and share highly conserved transcriptomes.

Lineage tracing is an important strategy for assessing the stem cell hierarchy as it allows stem and progenitor cell fate to be studied in situ in the context of development, tissue maintenance and disease. We have combined lineage tracing with a novel three-dimensional (3D) imaging strategy to explore the relative contributions of stem and progenitor cells to post-natal mammary gland development and tissue homeostasis. Cell lineage tracing studies also provide the current gold standard for identifying ‘cells of origin’ in cancer. Towards this end, we are utilizing newly generated transgenic strains harboring lineage-specific gene regulatory regions to direct the expression of specific mammary oncogenic lesions to discrete epithelial cell types. In terms of human breast cancer, the interrogation of molecular profiles has provided insight into potential ‘cells of origin’ in preneoplastic tissue for basal-like cancers arising in BRCA1 mutation carriers. Finally, the identification of deregulated genes/pathways in these crucial target cell populations may enable earlier detection of malignancies and lead to preventative therapies for individuals at high risk of developing breast cancer.

Citation Format: Jane E. Visvader, Anne Rios, Naiyang Fu, Emma Nolan, Geoffrey Lindeman. The breast epithelial hierarchy and its implications for tumor heterogeneity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr PL04-02. doi:10.1158/1538-7445.AM2014-PL04-02
Abstract SY12-04: Regulation of normal and cancer stem cell self renewal and senescence by USP16
Authors: Clarke Michael F.
Published: May 2015

Down syndrome (DS) is a complex clinical syndrome associated with multiple pathological conditions including deficits in motor skills and learning. DS has also been associated with early onset and higher incidence of aging-related phenomena such as Alzheimer's disease. Recently, it has been proposed that the aging process is correlated with an impaired or exhausted ability of stem cells to self-renew. we show that in Ts65Dn mice, trisomic for 132 genes homologous to HSA21, triplication of Usp16 reduces self-renewal of hematopoietic stem cells and expansion of mammary epithelial cells, neural progenitors, and fibroblasts. Moreover, Usp16 is associated with decreased ubiquitination of Cdkn2a and accelerated senescence in Ts65Dn fibroblasts. Usp16 can remove ubiquitin from H2AK119, a critical mark for the maintenance of multiple somatic tissues. Downregulation of Usp16, either by mutation of a single normal USP16 allele or by shRNAs, largely rescues all these defects. Furthermore, in human tissues overexpression of USP16 reduces the expansion of normal fibroblasts and post-natal neural progenitors while downregulation of USP16 partially rescues the proliferation defects of DS fibroblasts. Taken together, these results suggest that USP16 plays an important role in antagonizing the self-renewal and/or senescence pathways in Down syndrome and could serve as an attractive target to ameliorate some of the associated pathologies. Down syndrome patients also have a decreased incidence of solid tumors including breast cancer. The potential role of USP16 in human breast cancer will be discussed.

Citation Format: Michael F. Clarke. Regulation of normal and cancer stem cell self renewal and senescence by USP16. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr SY12-04. doi:10.1158/1538-7445.AM2014-SY12-04
Abstract SY20-04: Mechanisms of malignant progression
Authors: Weinberg Robert A.
Published: May 2015

Metastasis is responsible for ∼90% of cancer-associated mortality, but we still have an incomplete understanding of its underlying mechanisms. Thus, we don't understand why certain primary tumor cells have a predilection to disseminate while others do not; why disseminated cancer cells can thrive in certain distant tissues but not in others; what adaptations disseminated cancer cells originating in one type of primary tumor must undergo in order to thrive in the microenvironment of a distant and potentially inhospitable tissue; and why certain indolent disseminated micrometastatic deposits suddenly erupt after great delay, generating life-threatening disseminated metastatic diseases. These and related issues will be examined in this session.

Citation Format: Robert A. Weinberg. Mechanisms of malignant progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr SY20-04. doi:10.1158/1538-7445.AM2014-SY20-04
Abstract SY24-01: A SAGA of GCN5 and USP22 in stem cells and cancer
Authors: Dent Sharon Y. R. , Atanassov Boyko , Hirsch Calley , Koutelou Evangelia , Coban Zeynep , Lan Xian Jiang , Wang Li
Published: May 2015

Histone modifying enzymes are important creating and maintaining epigenetic programs that regulate cell identity and growth. Our lab uses genetic approaches to define the full spectrum of functions for these enzymes. For example, we have created a series of mutations in the mouse Gcn5 (KAT2A) gene in order to define the functions of this histone acetyltransferase (AT) in a mammalian system. Gcn5 is the catalytic subunit of the SAGA and ATAC complexes. Deletion of Gcn5 led to early embryonic death and to telomere dysfunction. We are now defining the role of Gcn5 and SAGA in maintenance of pluripotency in mouse ES cells and ES cell differentiation. Our data indicate that Gcn5 is an important cofactor for both Myc and E2F family transcription factors, predicting a role for Gcn5 not only in self renewal of ES cells but also in Myc-driven cancers. Our latest studies are defining the functions of the SAGA deubiquitinase module, and these are revealing new information about SAGA composition and function during mouse development and in human diseases.

Citation Format: Sharon Y. R. Dent, Boyko Atanassov, Calley Hirsch, Evangelia Koutelou, Zeynep Coban, Xian Jiang Lan, Li Wang. A SAGA of GCN5 and USP22 in stem cells and cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr SY24-01. doi:10.1158/1538-7445.AM2014-SY24-01